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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. [Effect of aspirin on function of human umbilical cord blood-derived late endothelial progenitor cells].

    PubMed

    Liu, Zhen-Zhen; Li, Guo-Qiang; Liu, Meng; Sun, Sheng-Xuan; An, Guan-Yu; Dong, Ning-Zheng

    2013-08-01

    This study was aimed to investigate whether aspirin has effect on function of late endothelial progenitor cells (EPC). Cord blood CD34(+) cells were purified using the ficoll density gradient centrifugation and human CD34 positive selection kit, then the cells were inoculated on fibronectin-coated culture plate. After culture for 2 weeks, adherent cells were identified as EPC by flow cytometry, immunofluorescence, RT-PCR, uptake of Dil-Ac-LDL and matrigel tube formation assay. EPC were treated with different concentrations of aspirin (0.1, 1, 10, 100, 1 000, 10 000 µmol/L) for 24 h, then the proliferation, adhesion and migration ability of these cells were analyzed by CCK-8 assay and transwell methods. The results indicated that the low concentrations of aspirin (0.1 and 1 000 µmol/L) promoted late EPC adhesive and migratory capacity, but no obvious effect on proliferation of late EPC were observed. On the other hand, the high concentrations of aspirin (10 000 µmol/L) inhibited proliferation and migratory capacity of EPC, but had no obvious effect on adhesive ability of EPC. It is concluded that low concentration of aspirin promotes migration and adhesion of late EPC, while the high concentration of aspirin decreases EPC proliferation and migratory capacity of EPC.

  3. Coculture of peripheral blood-derived mesenchymal stem cells and endothelial progenitor cells on strontium-doped calcium polyphosphate scaffolds to generate vascularized engineered bone.

    PubMed

    Fu, Wei-Li; Xiang, Zhou; Huang, Fu-Guo; Gu, Zhi-Peng; Yu, Xi-Xun; Cen, Shi-Qiang; Zhong, Gang; Duan, Xin; Liu, Ming

    2015-03-01

    Vascularization of engineered bone tissue is critical for ensuring its survival after implantation and it is the primary factor limiting its clinical use. A promising approach is to prevascularize bone grafts in vitro using endothelial progenitor cells (EPC) derived from peripheral blood. Typically, EPC are added together with mesenchymal stem cells (MSC) that differentiate into osteoblasts. One problem with this approach is how to promote traditional tissue engineering bone survival with a minimally invasive method. In this study, we examined the effectiveness of administering to stimulate the release of peripheral blood stem cells and their co-culturing system for generating prevascularized engineered bone. Cells were isolated by Ficoll density gradient centrifugation and identified as EPC and MSC based on morphology, surface markers, and functional analysis. EPC and MSC were cocultured in several different ratios, and cell morphology and tube formation were assessed by microscopy. Expression of osteogenesis and vascularization markers was quantified by enzyme-linked immunosorbent assay (ELISA), polymerase chain reaction, and histochemical and immunofluorescence staining. Increasing the proportion of EPC in the coculture system led to greater tube formation and greater expression of the endothelial cell marker CD31. An EPC:MSC ratio of 75:25 gave the highest expression of osteogenesis and angiogenesis markers. Cocultures adhered to a three-dimensional scaffold of strontium-doped calcium polyphosphate and proliferated well. Our findings show that coculturing peripheral blood-derived EPC and MSC may prove useful for generating prevascularized bone tissue for clinical use.

  4. Intravenous Administration of Human Umbilical Cord Blood-Derived AC133+ Endothelial Progenitor Cells in Rat Stroke Model Reduces Infarct Volume: Magnetic Resonance Imaging and Histological Findings

    PubMed Central

    Iskander, Asm; Knight, Robert A.; Zhang, Zheng Gang; Ewing, James R.; Shankar, Adarsh; Varma, Nadimpalli Ravi S.; Bagher-Ebadian, Hassan; Ali, Meser M.; Arbab, Ali S.

    2013-01-01

    Abstract Endothelial progenitor cells (EPCs) hold enormous therapeutic potential for ischemic vascular diseases. Previous studies have indicated that stem/progenitor cells derived from human umbilical cord blood (hUCB) improve functional recovery in stroke models. Here, we examined the effect of hUCB AC133+ EPCs on stroke development and resolution in a middle cerebral artery occlusion (MCAo) rat model. Since the success of cell therapies strongly depends on the ability to monitor in vivo the migration of transplanted cells, we also assessed the capacity of magnetic resonance imaging (MRI) to track in vivo the magnetically labeled cells that were administered. Animals were subjected to transient MCAo and 24 hours later injected intravenously with 107 hUCB AC133+ EPCs. MRI performed at days 1, 7, and 14 after the insult showed accumulation of transplanted cells in stroke-affected hemispheres and revealed that stroke volume decreased at a significantly higher rate in cell-treated animals. Immunohistochemistry analysis of brain tissues localized the administered cells in the stroke-affected hemispheres only and indicated that these cells may have significantly affected the magnitude of endogenous proliferation, angiogenesis, and neurogenesis. We conclude that transplanted cells selectively migrated to the ischemic brain parenchyma, where they exerted a therapeutic effect on the extent of tissue damage, regeneration, and time course of stroke resolution. PMID:23934909

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

    PubMed Central

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

    2015-01-01

    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. Equine peripheral blood-derived progenitors in comparison to bone marrow-derived mesenchymal stem cells.

    PubMed

    Koerner, Jens; Nesic, Dobrila; Romero, Jose Diaz; Brehm, Walter; Mainil-Varlet, Pierre; Grogan, Shawn Patrick

    2006-06-01

    Fibroblast-like cells isolated from peripheral blood of human, canine, guinea pig, and rat have been demonstrated to possess the capacity to differentiate into several mesenchymal lineages. The aim of this work was to investigate the possibility of isolating pluripotent precursor cells from equine peripheral blood and compare them with equine bone marrow-derived mesenchymal stem cells. Human mesenchymal stem cells (MSCs) were used as a control for cell multipotency assessment. Venous blood (n = 33) and bone marrow (n = 5) were obtained from adult horses. Mononuclear cells were obtained by Ficoll gradient centrifugation and cultured in monolayer, and adherent fibroblast-like cells were tested for their differentiation potential. Chondrogenic differentiation was performed in serum-free medium in pellet cultures as a three-dimensional model, whereas osteogenic and adipogenic differentiation were induced in monolayer culture. Evidence for differentiation was made via biochemical, histological, and reverse transcription-polymerase chain reaction evaluations. Fibroblast-like cells were observed on day 10 in 12 out of 33 samples and were allowed to proliferate until confluence. Equine peripheral blood-derived cells had osteogenic and adipogenic differentiation capacities comparable to cells derived from bone marrow. Both cell types showed a limited capacity to produce lipid droplets compared to human MSCs. This result may be due to the assay conditions, which are established for human MSCs from bone marrow and may not be optimal for equine progenitor cells. Bone marrow-derived equine and human MSCs could be induced to develop cartilage, whereas equine peripheral blood progenitors did not show any capacity to produce cartilage at the histological level. In conclusion, equine peripheral blood-derived fibroblast-like cells can differentiate into distinct mesenchymal lineages but have less multipotency than bone marrow-derived MSCs under the conditions used in this study.

  7. Circulating and tissue resident endothelial progenitor cells.

    PubMed

    Basile, David P; Yoder, Mervin C

    2014-01-01

    Progenitor cells for the endothelial lineage have been widely investigated for more than a decade, but continue to be controversial since no unique identifying marker has yet been identified. This review will begin with a discussion of the basic tenets originally proposed for proof that a cell displays properties of an endothelial progenitor cell. We then provide an overview of the methods for putative endothelial progenitor cell derivation, expansion, and enumeration. This discussion includes consideration of cells that are present in the circulation as well as cells resident in the vascular endothelial intima. Finally, we provide some suggested changes in nomenclature that would greatly clarify and demystify the cellular elements involved in vascular repair.

  8. Endothelial progenitor cells in cardiovascular diseases.

    PubMed

    Lee, Poay Sian Sabrina; Poh, Kian Keong

    2014-07-26

    Endothelial dysfunction has been associated with the development of atherosclerosis and cardiovascular diseases. Adult endothelial progenitor cells (EPCs) are derived from hematopoietic stem cells and are capable of forming new blood vessels through a process of vasculogenesis. There are studies which report correlations between circulating EPCs and cardiovascular risk factors. There are also studies on how pharmacotherapies may influence levels of circulating EPCs. In this review, we discuss the potential role of endothelial progenitor cells as both diagnostic and prognostic biomarkers. In addition, we look at the interaction between cardiovascular pharmacotherapies and endothelial progenitor cells. We also discuss how EPCs can be used directly and indirectly as a therapeutic agent. Finally, we evaluate the challenges facing EPC research and how these may be overcome.

  9. 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

  10. 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.

  11. Endothelial progenitor cells--an evolving story.

    PubMed

    Pearson, Jeremy D

    2010-05-01

    The first description of endothelial progenitor cells (EPC) in 1997 led rapidly to substantial changes in our understanding of angiogenesis, and within 5 years to the first clinical studies in humans using bone marrow derived EPC to enhance coronary neovascularisation and cardiac function after myocardial ischemia. However, to improve the success of this therapy a clearer understanding of the biology of EPC is needed. This article summarises recent data indicating that most EPC are not, in fact, endothelial progenitors but can be better described as angiogenic monocytes, and explores the implications this has for their future therapeutic use.

  12. 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

  13. 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

  14. Endothelial progenitor cells in diabetic foot syndrome.

    PubMed

    Drela, Ewelina; Stankowska, Katarzyna; Kulwas, Arleta; Rość, Danuta

    2012-01-01

    In the late 20th century endothelial progenitor cells (EPCs) were discovered and identified as cells capable of differentiating into endothelial cells. Antigens characteristic of endothelial cells and hematopoietic cells are located on their surface. EPCs can proliferate, adhere, migrate and have the specific ability to form vascular structure, and they have a wide range of roles: They participate in maintaining hemostasis, and play an important part in the processes of vasculogenesis and angiogenesis. They are sources of angiogenic factors, especially vascular endothelial growth factor (VEGF). EPCs exist in bone marrow, from which they are recruited into circulation in response to specific stimuli. Tissue ischemia is thought to be the strongest inductor of EPC mobilization. Local ischemia accompanies many pathological states, including diabetic foot syndrome (DFS). Impaired angiogenesis--in which EPCs participate--is typical of DFS. An analysis of the available literature indicates that in diabetic patients the number of EPCs declines and their functioning is impaired. Endothelial progenitor cells are crucial to vasculogenesis and angiogenesis during ischemic neovascularization. The pathomechanisms underlying impaired angiogenesis in patients with DFS is complicated, but the discovery of EPCs has shed new light on the pathogenesis of many diseases, including diabetes foot syndrome.

  15. Enhancing endothelial progenitor cell for clinical use

    PubMed Central

    Ye, Lei; Poh, Kian-Keong

    2015-01-01

    Circulating endothelial progenitor cells (EPCs) have been demonstrated to correlate negatively with vascular endothelial dysfunction and cardiovascular risk factors. However, translation of basic research into the clinical practice has been limited by the lack of unambiguous and consistent definitions of EPCs and reduced EPC cell number and function in subjects requiring them for clinical use. This article critically reviews the definition of EPCs based on commonly used protocols, their value as a biomarker of cardiovascular risk factor in subjects with cardiovascular disease, and strategies to enhance EPCs for treatment of ischemic diseases. PMID:26240678

  16. Enhancing endothelial progenitor cell for clinical use.

    PubMed

    Ye, Lei; Poh, Kian-Keong

    2015-07-26

    Circulating endothelial progenitor cells (EPCs) have been demonstrated to correlate negatively with vascular endothelial dysfunction and cardiovascular risk factors. However, translation of basic research into the clinical practice has been limited by the lack of unambiguous and consistent definitions of EPCs and reduced EPC cell number and function in subjects requiring them for clinical use. This article critically reviews the definition of EPCs based on commonly used protocols, their value as a biomarker of cardiovascular risk factor in subjects with cardiovascular disease, and strategies to enhance EPCs for treatment of ischemic diseases.

  17. Endothelial progenitor cell dysfunction in rheumatic disease.

    PubMed

    Westerweel, Peter E; Verhaar, Marianne C

    2009-06-01

    Rheumatic disease is characterized by inflammation and endothelial dysfunction, which contribute to accelerated atherosclerosis. Circulating endothelial progenitor cells (EPCs) can restore dysfunctional endothelium and thereby protect against atherosclerotic vascular disease. The number and function of EPCs are, however, affected in rheumatic diseases such as psoriatic arthritis, rheumatoid arthritis, systemic lupus erythematosus, systemic sclerosis, and antineutrophil cytoplasmic autoantibody-associated vasculitis. rheumatic disease is often characterized by decreased numbers, and impaired function, of EPCs, although numbers of these cells might increase during the initial years of systemic sclerosis. Pioneering studies show that EPC dysfunction might be improved with pharmacological treatment. How best to restore EPC function, and whether achieving this aim can prevent long-term cardiovascular complications in rheumatic disease, remain to be established.

  18. 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

  19. Endothelial progenitor cells: a new player in lupus?

    PubMed

    Haque, Sahena; Alexander, M Yvonne; Bruce, Ian N

    2012-01-01

    Patients with systemic lupus erythematosus (SLE) have a greatly increased risk of cardiovascular disease. There is growing interest in the link between vascular damage and lupus-specific inflammatory factors. Impaired endothelial repair could account for the endothelial dysfunction in this patient group. This review describes the contribution that endothelial progenitor cells could play in the pathogenesis of premature vascular damage in this disease. The methods of isolation, detection, and characterization of endothelial progenitor cells, together with their potential role in repair of the endothelium and as a therapeutic target in SLE, are discussed. PMID:22356717

  20. Toll-like receptor 3 regulates cord blood-derived endothelial cell function in vitro and in vivo.

    PubMed

    Grelier, Aurore; Cras, Audrey; Balitrand, Nicole; Delmau, Catherine; Lecourt, Séverine; Lepelletier, Yves; Riesterer, Hélène; Freida, Delphine; Lataillade, Jean-Jacques; Lebousse-Kerdiles, Marie-Caroline; Cuccini, Wendy; Peffault de Latour, Regis; Marolleau, Jean-Pierre; Uzan, Georges; Larghero, Jérôme; Vanneaux, Valérie

    2013-10-01

    Circulating endothelial progenitor cells (cEPC) are capable of homing to neovascularisation sites, in which they proliferate and differentiate into endothelial cells. Transplantation of cEPC-derived cells, in particular those isolated from umbilical cord blood (UCB), has emerged as a promising approach in the treatment of cardio-vascular diseases. After in vivo transplantation, these cells may be exposed to local or systemic inflammation or pathogens, of which they are a common target. Because Toll-like receptors (TLR) are critical in detecting pathogens and in initiating inflammatory responses, we hypothesized that TLR may govern UCB cEPC-derived cells function. While these cells expressed almost all TLR, we found that only TLR3 dramatically impaired cell properties. TLR3 activation inhibited cell proliferation, modified cell cycle entry, impaired the in vitro angiogenic properties and induced pro-inflammatory cytokines production. The anti-angiogenic effect of TLR3 activation was confirmed in vivo in a hind-limb ischemic mice model. Moreover, TLR3 activation consistently leads to an upregulation of miR-29b, -146a and -155 and to a deregulation of cytoskeleton and cell cycle regulator. Hence, TLR3 activation is likely to be a key regulator of cEPC-derived cells properties.

  1. 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.

  2. 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

  3. 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

  4. 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

  5. Development of Endothelial-Specific Single Inducible Lentiviral Vectors for Genetic Engineering of Endothelial Progenitor Cells.

    PubMed

    Yang, Guanghua; Kramer, M Gabriela; Fernandez-Ruiz, Veronica; Kawa, Milosz P; Huang, Xin; Liu, Zhongmin; Prieto, Jesus; Qian, Cheng

    2015-11-27

    Endothelial progenitor cells (EPC) are able to migrate to tumor vasculature. These cells, if genetically modified, can be used as vehicles to deliver toxic material to, or express anticancer proteins in tumor. To test this hypothesis, we developed several single, endothelial-specific, and doxycycline-inducible self-inactivating (SIN) lentiviral vectors. Two distinct expression cassettes were inserted into a SIN-vector: one controlled by an endothelial lineage-specific, murine vascular endothelial cadherin (mVEcad) promoter for the expression of a transactivator, rtTA2S-M2; and the other driven by an inducible promoter, TREalb, for a firefly luciferase reporter gene. We compared the expression levels of luciferase in different vector constructs, containing either the same or opposite orientation with respect to the vector sequence. The results showed that the vector with these two expression cassettes placed in opposite directions was optimal, characterized by a robust induction of the transgene expression (17.7- to 73-fold) in the presence of doxycycline in several endothelial cell lines, but without leakiness when uninduced. In conclusion, an endothelial lineage-specific single inducible SIN lentiviral vector has been developed. Such a lentiviral vector can be used to endow endothelial progenitor cells with anti-tumor properties.

  6. 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

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

    PubMed

    Aragona, Caterina Oriana; Imbalzano, Egidio; Mamone, Federica; Cairo, Valentina; Lo Gullo, Alberto; D'Ascola, Angela; Sardo, Maria Adriana; Scuruchi, Michele; Basile, Giorgio; Saitta, Antonino; 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.

  8. 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

  9. 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

  10. 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

  11. 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

  12. Isolation and angiogenesis by endothelial progenitors in the fetal liver.

    PubMed

    Cherqui, Stephanie; Kurian, Sunil M; Schussler, Olivier; Hewel, Johannes A; Yates, John R; Salomon, Daniel R

    2006-01-01

    Endothelial progenitor cells (EPCs) have significant therapeutic potential. However, the low quantity of such cells available from bone marrow and their limited capacity to proliferate in culture make their use difficult. Here, we present the first definitive demonstration of the presence of true EPCs in murine fetal liver capable of forming blood vessels in vivo connected to the host's vasculature after transplantation. This population is particularly interesting because it can be obtained at high yield and has a high angiogenic capacity as compared with bone marrow-derived EPCs. The EPC capacity is contained within the CD31+Sca1+ cell subset. We demonstrate that these cells are dependent for survival and proliferation on a feeder cell monolayer derived from the fetal liver. In addition, we describe a novel and easy method for the isolation and ex vivo proliferation of these EPCs. Finally, we used gene expression profiling and tandem mass spectrometry proteomics to examine the fetal liver endothelial progenitors and the feeder cells to identify possible proangiogenic growth factor and endothelial differentiation-associated genes.

  13. Isolation and characterization of endothelial progenitor cells from human blood.

    PubMed

    Mead, Laura E; Prater, Daniel; Yoder, Mervin C; Ingram, David A

    2008-07-01

    Circulating endothelial progenitor cells (EPCs) in adult human peripheral blood were originally identified in 1997 by Asahara et al., which challenged the paradigm that vasculogenesis is a process restricted to embryonic development. Since their original identification, EPCs have been extensively studied as biomarkers to assess the risk of cardiovascular disease in human subjects and as a potential cell therapeutic for vascular regeneration. Endothelial colony-forming cells (ECFCs), which are a subtype of EPCs, were recently identified from circulating adult and human umbilical cord blood. In contrast to other types of EPCs, which display various monocyte/macrophage phenotypes and functions, ECFCs are characterized by robust proliferative potential, secondary and tertiary colony formation upon replating, and de novo blood vessel formation in vivo when transplanted into immunodeficient mice. In this unit, we describe detailed methodologies for isolation and characterization of ECFCs from both human peripheral and umbilical cord blood.

  14. Early endothelial progenitor cells as a source of myeloid cells to improve the pre-vascularisation of bone constructs.

    PubMed

    Shi, Y; Kramer, G; Schröder, A; Kirkpatrick, C J; Seekamp, A; Schmidt, H; Fuchs, S

    2014-01-25

    According to present knowledge, blood derived endothelial progenitor cells (EPC) might act as proangiogenic myeloid cells, which play a fundamental role in the regulation of angiogenesis and blood vessel reorganisation. In this context, we have evaluated the contribution of endogenous myeloid cells in co-cultures of blood derived outgrowth endothelial cells (OEC) and osteogenic cells. In addition, we investigated the role of EPC as a potential source of myeloid cells in the formation of vascular structures in an in vitro model consisting of mesenchymal stem cells (MSC) and OEC. For this purpose, we added EPCs to co-cultures of MSC and OECs. Vascular structures and the co-localisation of myeloid cells were analysed by confocal laser microscopy (CLSM) for endothelial and myeloid markers and quantitative image analysis. The molecular effects of myeloid cells were evaluated by quantitative real time PCR, ELISA and protein arrays from cell culture supernatants and lysates. Endogenous myeloid cells were significantly co-localised with angiogenic structures in co-cultures of OEC and osteogenic cells. The active addition of EPC to co-cultures of OEC and MSC resulted in a statistically approved increase in the formation of prevascular structures at early stages of the co-culture process. In addition, we observed an increase of endothelial markers, indicating beneficial effects of EPC or myeloid cells on endothelial cell growth. Furthermore, real time PCR indicated high expression levels of CD68, CD11b and CD163 in co-cultures of EPC and MSC indicating that EPC act at least partly as macrophage like-cells.

  15. 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

  16. Extrarenal Progenitor Cells Do Not Contribute to Renal Endothelial Repair.

    PubMed

    Sradnick, Jan; Rong, Song; Luedemann, Anika; Parmentier, Simon P; Bartaun, Christoph; Todorov, Vladimir T; Gueler, Faikah; Hugo, Christian P; Hohenstein, Bernd

    2016-06-01

    Endothelial progenitor cells (EPCs) may be relevant contributors to endothelial cell (EC) repair in various organ systems. In this study, we investigated the potential role of EPCs in renal EC repair. We analyzed the major EPC subtypes in murine kidneys, blood, and spleens after induction of selective EC injury using the concanavalin A/anti-concanavalin A model and after ischemia/reperfusion (I/R) injury as well as the potential of extrarenal cells to substitute for injured local EC. Bone marrow transplantation (BMTx), kidney transplantation, or a combination of both were performed before EC injury to allow distinction of extrarenal or BM-derived cells from intrinsic renal cells. During endothelial regeneration, cells expressing markers of endothelial colony-forming cells (ECFCs) were the most abundant EPC subtype in kidneys, but were not detected in blood or spleen. Few cells expressing markers of EC colony-forming units (EC-CFUs) were detected. In BM chimeric mice (C57BL/6 with tandem dimer Tomato-positive [tdT+] BM cells), circulating and splenic EC-CFUs were BM-derived (tdT+), whereas cells positive for ECFC markers in kidneys were not. Indeed, most BM-derived tdT+ cells in injured kidneys were inflammatory cells. Kidneys from C57BL/6 donors transplanted into tdT+ recipients with or without prior BMTx from C57BL/6 mice were negative for BM-derived or extrarenal ECFCs. Overall, extrarenal cells did not substitute for any intrinsic ECs. These results demonstrate that endothelial repair in mouse kidneys with acute endothelial lesions depends exclusively on local mechanisms.

  17. Association of SIRT1 expression with shear stress induced endothelial progenitor cell differentiation.

    PubMed

    Cheng, Bin-Bin; Yan, Zhi-Qiang; Yao, Qing-Ping; Shen, Bao-Rong; Wang, Ji-Yao; Gao, Li-Zhi; Li, Yu-Qing; Yuan, Hai-Tao; Qi, Ying-Xin; Jiang, Zong-Lai

    2012-12-01

    Shear stress imposed by blood flow is crucial for differentiation of endothelial progenitor cells (EPCs). Histone deacetylase SIRT1 has been shown to play a pivotal role in many physiological processes. However, association of SIRT1 expression with shear stress-induced EPC differentiation remains to be elucidated. The present study was designed to determine the effect of SIRT1 on EPC differentiation induced by shear stress, and to seek the underlying mechanisms. Human umbilical cord blood-derived EPCs were exposed to laminar shear stress of 15 dyn/cm(2) by parallel plate flow chamber system. Shear stress enhanced EPC differentiation toward endothelial cells (ECs) while inhibited to smooth muscle cells (SMCs). The expressions of phospho-Akt, SIRT1 and histone H3 acetylation (Ac-H3) in EPCs were detected after exposure to shear stress for 2, 6, 12, and 24 h, respectively. Shear stress significantly activated Akt phosphorylation, augmented SIRT1 expression and downregulated Ac-H3. SIRT1 siRNA in EPCs diminished the expression of EC markers, but increased the expression of SMC markers, and resulted in upregulation of Ac-H3. Whereas, resveratrol, an activator of SIRT1, had the opposite effects on both EPC differentiation and histone H3 acetylation. Wortmannin, an inhibitor of PI3-kinase, suppressed endothelial differentiation of EPCs, decreased SIRT1, and upregulated Ac-H3 expression. In addition, SIRT1 promoted tube formation of EPCs in matrix gels. These results provided a mechanobiological basis of shear stress-induced EPC differentiation into ECs and suggest that PI3k/Akt-SIRT1-Ac-H3 pathway is crucial in such a process.

  18. 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

  19. 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

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

    PubMed Central

    Capriglione, Luiz Guilherme A; Barchiki, Fabiane; Miyague, Lye; Jackowski, Danielle; Fracaro, Letícia; Schittini, Andressa V; Senegaglia, Alexandra C; Rebelatto, Carmen LK; Olandoski, Márcia; Correa, Alejandro; Brofman, Paulo RS

    2015-01-01

    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 cell recruitment in a microfluidic vascular model.

    PubMed

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

    2015-01-01

    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

  2. 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

  3. Cilengitide inhibits proliferation and differentiation of human endothelial progenitor cells in vitro

    SciTech Connect

    Loges, Sonja; Butzal, Martin; Otten, Jasmin; Schweizer, Michaela; Fischer, Uta; Bokemeyer, Carsten; Hossfeld, Dieter K.; Schuch, Gunter; Fiedler, Walter . E-mail: fiedler@uke.uni-hamburg.de

    2007-06-15

    Bone marrow derived hematopoietic stem cells can function as endothelial progenitor cells. They are recruited to malignant tumors and differentiate into endothelial cells. This mechanism of neovascularization termed vasculogenesis is distinct from proliferation of pre-existing vessels. To better understand vasculogenesis we developed a cell culture model with expansion and subsequent endothelial differentiation of human CD133{sup +} progenitor cells in vitro. {alpha}{sub v}{beta}{sub 3}-integrins are expressed by endothelial cells and play a role in the attachment of endothelial cells to the extracellular matrix. We investigated the effect of Cilengitide, a peptide-like, high affinity inhibitor of {alpha}{sub v}{beta}{sub 3}- and {alpha}{sub v}{beta}{sub 5}-integrins in our in vitro system. We could show expression of {alpha}{sub v}{beta}{sub 3}-integrin on 60 {+-} 9% of non-adherent endothelial progenitors and on 91 {+-} 7% of differentiated endothelial cells. {alpha}{sub v}{beta}{sub 3}-integrin was absent on CD133{sup +} hematopoietic stem cells. Cilengitide inhibited proliferation of CD133{sup +} cells in a dose-dependent manner. The development of adherent endothelial cells from expanded CD133{sup +} cells was reduced even stronger by Cilengitide underlining its effect on integrin mediated cell adhesion. Expression of endothelial antigens CD144 and von Willebrand factor on differentiating endothelial precursors was decreased by Cilengitide. In summary, Cilengitide inhibits proliferation and differentiation of human endothelial precursor cells underlining its anti-angiogenic effects.

  4. Oxidized low-density lipoprotein alters endothelial progenitor cell populations.

    PubMed

    Cui, Yuqi; Narasimhulu, Chandrakala A; Liu, Lingjuan; Li, Xin; Xiao, Yuan; Zhang, Jia; Xie, Xiaoyun; Hao, Hong; Liu, Jason Z; He, Guanglong; Cowan, Peter J; Cui, Lianqun; Zhu, Hua; Parthasarathy, Sampath; Liu, Zhenguo

    2015-06-01

    Oxidized low-density lipoprotein (ox-LDL) is critical to atherosclerosis in hyperlipidemia. Bone marrow (BM)-derived endothelial progenitor cells (EPCs) are important to preventing atherosclerosis, and significantly decreased in hyperlipidemia. This study was to demonstrate ox-LDL and hyperlipidemia could exhibit similar effect on EPC population and the role of reactive oxygen species (ROS). ROS production in BM and blood was significantly increased in male C57BL/6 mice with intravenous ox-LDL treatment, and in hyperlipidemic LDL receptor knockout mice with 4-month high-fat diet. ROS formation was effectively blocked with overexpression of antioxidant enzymes or N-acetylcysteine treatment. In hyperlipidemic and ox-LDL-treated mice, c-Kit(+)/CD31(+) cell number in BM and blood, and Sca-1(+)/Flk-1(+) cell number in blood, not in BM, were significantly decreased, which were not affected by inhibiting ROS production, while blood CD34(+)/Flk-1(+) cell number was significantly increased that was prevented with reduced ROS formation. However, blood CD34(+)/CD133(+) cell number increased in ox-LDL-treated mice, while decreased in hyperlipidemic mice. These data suggested that ox-LDL produced significant changes in BM and blood EPC populations similar (but not identical) to chronic hyperlipidemia with predominantly ROS-independent mechanism(s).

  5. 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

  6. 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

  7. 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

  8. Impaired function of bone marrow-derived endothelial progenitor cells in murine liver fibrosis.

    PubMed

    Shirakura, Katsuya; Masuda, Haruchika; Kwon, Sang-Mo; Obi, Syotaro; Ito, Rie; Shizuno, Tomoko; Kurihara, Yusuke; Mine, Tetsuya; Asahara, Takayuki

    2011-01-01

    Liver fibrosis (LF) caused by chronic liver damage has been considered as an irreversible disease. As alternative therapy for liver transplantation, there are high expectations for regenerative medicine of the liver. Bone marrow (BM)- or peripheral blood-derived stem cells, including endothelial progenitor cells (EPCs), have recently been used to treat liver cirrhosis. We investigated the biology of BM-derived EPC in a mouse model of LF. C57BL/6J mice were subcutaneously injected with carbon tetrachloride (CCl(4)) every 3 days for 90 days. Sacrificed 2 days after final injection, whole blood (WB) was collected for isolation of mononuclear cells (MNCs) and biochemical examination. Assessments of EPC in the peripheral blood and BM were performed by flow cytometry and EPC colony-forming assay, respectively, using purified MNCs and BM c-KIT(+), Sca-1(+), and Lin(-) (KSL) cells. Liver tissues underwent histological analysis with hematoxylin/eosin/Azan staining, and spleens were excised and weighed. CCl(4)-treated mice exhibited histologically bridging fibrosis, pseudolobular formation, and splenomegaly, indicating successful induction of LF. The frequency of definitive EPC-colony-forming-units (CFU) as well as total EPC-CFU at the equivalent cell number of 500 BM-KSL cells decreased significantly (p < 0.0001) in LF mice compared with control mice; no significant changes in primitive EPC-CFU occurred in LF mice. The frequency of WB-MNCs of definitive EPC-CFU decreased significantly (p < 0.01) in LF mice compared with control mice. Together, these findings indicated the existence of impaired EPC function and differentiation in BM-derived EPCs in LF mice and might be related to clinical LF.

  9. 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

  10. 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

  11. 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

  12. 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

  13. Glycogen synthase kinase 3β inhibition enhanced proliferation, migration and functional re-endothelialization of endothelial progenitor cells in hypercholesterolemia microenvironment

    PubMed Central

    Cui, Bin; Jin, Jun; Ding, Xiaohan; Deng, Mengyang; Yu, Shiyong; Song, MingBao; Yu, Yang; Zhao, Xiaohui; Chen, Jianfei

    2015-01-01

    Hypercholesterolemia impairs the quantity and function of endothelial progenitor cell. We hypothesized that glycogen synthase kinase 3β activity is involved in regulating biological function of endothelial progenitor cells in hypercholesterolemia microenvironment. For study, endothelial progenitor cells derived from apolipoprotein E-deficient mice fed with high-fat diet were used. Glycogen synthase kinase 3β activity was interfered with glycogen synthase kinase 3β inhibitor lithium chloride or transduced with replication defective adenovirus vector expressing catalytically inactive glycogen synthase kinase 3β (GSK3β-KM). Functions of endothelial progenitor cells, proliferation, migration, secretion and network formation of endothelial progenitor cells were assessed in vitro. The expression of phospho-glycogen synthase kinase 3β, β-catenin and cyclinD1 in endothelial progenitor cells was detected by Western blot. The in vivo function re-endothelialization and vasodilation were also analyzed by artery injury model transplanted with glycogen synthase kinase 3β-inhibited endothelial progenitor cells. We demonstrated that while the proliferation, migration, network formation as well as VEGF and NO secretion were impaired in apolipoprotein E-deficient endothelial progenitor cells, glycogen synthase kinase 3β inhibition significantly improved all these functions. Apolipoprotein E-deficient endothelial progenitor cells showed decreased phospho-glycogen synthase kinase 3β, β-catenin and cyclinD1 expression, whereas these signals were enhanced by glycogen synthase kinase 3β inhibition and accompanied with β-catenin nuclear translocation. Our in vivo model showed that glycogen synthase kinase 3β inhibition remarkably increased re-endothelial and vasodilation. Taken together, our data suggest that inhibition of glycogen synthase kinase 3β is associated with endothelial progenitor cell biological functions both in vitro and in vivo. It might be an important

  14. 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

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

    PubMed

    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.

  16. Conditioned medium from human umbilical vein endothelial cells markedly improves the proliferation and differentiation of circulating endothelial progenitors.

    PubMed

    Castelli, Germana; Parolini, Isabella; Cerio, Anna Maria; D'Angiò, Agnese; Pasquini, Luca; Carollo, Maria; Sargiacomo, Massimo; Testa, Ugo; Pelosi, Elvira

    2016-10-01

    Circulating endothelial progenitor cells (EPCs) have been suggested as a precious source for generating functionally competent endothelial cells (ECs), candidate for various clinical applications. However, the paucity of these progenitor cells and the technical difficulties for their in vitro growth represent a main limitation to their use. In the present study we hypothesized that the paracrine effects of human umbilical vein endothelial cells (HUVECs) may improve endothelial cell generation from cord blood (CB) EPCs. In line with this hypothesis we showed that HUVEC conditioned medium (CM) or co-culture with HUVECs markedly improved the proliferation and differentiation and delayed the senescence of CB EPCs. The endothelial-promoting effect of CM seems to be related to smaller vesicles including exosomes (sEV/exo) contained in this medium and transferred to CB CD34(+) EPCs: in fact, purified preparations of sEV/exo isolated from CM mimicked the effect of CM to sustain endothelial formation. These observations provided the interesting indication that mature ECs exert a stimulatory effect on endothelial cell differentiation from CD34(+) cells. PMID:27667168

  17. Human endothelial progenitor cells internalize high-density lipoprotein.

    PubMed

    Srisen, Kaemisa; Röhrl, Clemens; Meisslitzer-Ruppitsch, Claudia; Ranftler, Carmen; Ellinger, Adolf; Pavelka, Margit; Neumüller, Josef

    2013-01-01

    Endothelial progenitor cells (EPCs) originate either directly from hematopoietic stem cells or from a subpopulation of monocytes. Controversial views about intracellular lipid traffic prompted us to analyze the uptake of human high density lipoprotein (HDL), and HDL-cholesterol in human monocytic EPCs. Fluorescence and electron microscopy were used to investigate distribution and intracellular trafficking of HDL and its associated cholesterol using fluorescent surrogates (bodipy-cholesterol and bodipy-cholesteryl oleate), cytochemical labels and fluorochromes including horseradish peroxidase and Alexa Fluor® 568. Uptake and intracellular transport of HDL were demonstrated after internalization periods from 0.5 to 4 hours. In case of HDL-Alexa Fluor® 568, bodipy-cholesterol and bodipy-cholesteryl oleate, a photooxidation method was carried out. HDL-specific reaction products were present in invaginations of the plasma membrane at each time of treatment within endocytic vesicles, in multivesicular bodies and at longer periods of uptake, also in lysosomes. Some HDL-positive endosomes were arranged in form of "strings of pearl"- like structures. HDL-positive multivesicular bodies exhibited intensive staining of limiting and vesicular membranes. Multivesicular bodies of HDL-Alexa Fluor® 568-treated EPCs showed multilamellar intra-vacuolar membranes. At all periods of treatment, labeled endocytic vesicles and organelles were apparent close to the cell surface and in perinuclear areas around the Golgi apparatus. No HDL-related particles could be demonstrated close to its cisterns. Electron tomographic reconstructions showed an accumulation of HDL-containing endosomes close to the trans-Golgi-network. HDL-derived bodipy-cholesterol was localized in endosomal vesicles, multivesicular bodies, lysosomes and in many of the stacked Golgi cisternae and the trans-Golgi-network Internalized HDL-derived bodipy-cholesteryl oleate was channeled into the lysosomal intraellular

  18. Adhesion behavior of endothelial progenitor cells to endothelial cells in simple shear flow

    NASA Astrophysics Data System (ADS)

    Gong, Xiao-Bo; Li, Yu-Qing; Gao, Quan-Chao; Cheng, Bin-Bin; Shen, Bao-Rong; Yan, Zhi-Qiang; Jiang, Zong-Lai

    2011-12-01

    The adhesion of endothelial progenitor cells (EPCs) on endothelial cells (ECs) is one of the critical physiological processes for the regenesis of vascular vessels and the prevention of serious cardiovascular diseases. Here, the rolling and adhesion behavior of EPCs on ECs was studied numerically. A two-dimensional numerical model was developed based on the immersed boundary method for simulating the rolling and adhesion of cells in a channel flow. The binding force arising from the catch bond of a receptor and ligand pair was modeled with stochastic Monte Carlo method and Hookean spring model. The effect of tumor necrosis factor alpha (TNF- α) on the expression of the number of adhesion molecules in ECs was analyzed experimentally. A flow chamber system with CCD camera was set up to observe the top view of the rolling of EPCs on the substrate cultivated with ECs. Numerical results prove that the adhesion of EPC on ECs is closely related to membrane stiffness of the cell and shear rate of the flow. It also suggests that the adhesion force between EPC and EC by P-selectin glycoprotein ligand-1 only is not strong enough to bond the cell onto vessel walls unless contributions of other catch bond are considered. Experimental results demonstrate that TNF- α enhanced the expressions of VCAM, ICAM, P-selectin and E-selectin in ECs, which supports the numerical results that the rolling velocity of EPC on TNF- α treated EC substrate decreases obviously compared with its velocity on the untreated one. It is found that because the adhesion is affected by both the rolling velocity and the deformability of the cell, an optimal stiffness of EPC may exist at a given shear rate of flow for achieving maximum adhesion rates.

  19. 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

  20. 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.

  1. Dysfunctional endothelial progenitor cells in cardiovascular diseases: role of NADPH oxidase.

    PubMed

    Peng, Jun; Liu, Bin; Ma, Qi-Lin; Luo, Xiu-Ju

    2015-01-01

    Endothelial progenitor cells (EPCs) play a critical role in maintenance of the endothelial integrity and vascular homeostasis, as well as in neovascularization. Dysfunctional EPCs are believed to contribute to the endothelial dysfunction and are closely related to the development of various cardiovascular diseases, such as hypertension, hyperlipidemia, and stroke. However, the underlying mechanisms of EPC dysfunction are complicated and remain largely elusive. Recent studies have demonstrated that reactive oxygen species (ROS) are key factors that involve in modulation of stem and progenitor cell function under various physiologic and pathologic conditions. It has been shown that NADPH oxidase (NOX)-derived ROS are the major sources of ROS in cardiovascular system. Accumulating evidence suggests that NOX-mediated oxidative stress can modulate EPC bioactivities, such as mobilization, migration, and neovascularization, and that inhibition of NOX has been shown to improve EPC functions. This review summarized recent progress in the studies on the correlation between NOX-mediated EPC dysfunction and cardiovascular diseases.

  2. 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

  3. Improving cardiovascular outcomes in rheumatic diseases: therapeutic potential of circulating endothelial progenitor cells.

    PubMed

    Reynolds, John A; Robertson, Abigail C; Bruce, Ian N; Alexander, M Yvonne

    2014-05-01

    Patients with Rheumatoid Arthritis (RA) and Systemic Lupus Erythematosus (SLE) have a significantly increased risk of cardiovascular disease (CVD). The reason for this is unclear but may be due, at least in part, to the failure of endothelial repair mechanisms. Over the last 15 years there has been much interest in the mechanisms of endothelial renewal and its potential as a therapy for CVD. In the circulation there are two distinct populations of cells; myeloid angiogenic cells (MACs) which augment repair by the paracrine secretion of angiogenic factors, and outgrowth endothelial cells (OECs) which are true endothelial progenitor cells (EPCs) and promote vasculogenesis by differentiating into mature endothelium. There are marked abnormalities in the number and function of these cells in patients with RA and SLE. Inflammatory cytokines including interferon-alpha (IFNα) and tumour-necrosis factor alpha (TNFα) both impair MAC and OEC function ex vivo and may therefore contribute to the CVD risk in these patients. Whilst administration of mononuclear cells, MACs and other progenitors has improved cardiovascular outcomes in the acute setting, this is not a viable option in chronic disease. The pharmacological manipulation of MAC/OEC function in vivo however has the potential to significantly improve endothelial repair and thus reduce CVD in this high risk population. PMID:24333265

  4. Improving cardiovascular outcomes in rheumatic diseases: therapeutic potential of circulating endothelial progenitor cells.

    PubMed

    Reynolds, John A; Robertson, Abigail C; Bruce, Ian N; Alexander, M Yvonne

    2014-05-01

    Patients with Rheumatoid Arthritis (RA) and Systemic Lupus Erythematosus (SLE) have a significantly increased risk of cardiovascular disease (CVD). The reason for this is unclear but may be due, at least in part, to the failure of endothelial repair mechanisms. Over the last 15 years there has been much interest in the mechanisms of endothelial renewal and its potential as a therapy for CVD. In the circulation there are two distinct populations of cells; myeloid angiogenic cells (MACs) which augment repair by the paracrine secretion of angiogenic factors, and outgrowth endothelial cells (OECs) which are true endothelial progenitor cells (EPCs) and promote vasculogenesis by differentiating into mature endothelium. There are marked abnormalities in the number and function of these cells in patients with RA and SLE. Inflammatory cytokines including interferon-alpha (IFNα) and tumour-necrosis factor alpha (TNFα) both impair MAC and OEC function ex vivo and may therefore contribute to the CVD risk in these patients. Whilst administration of mononuclear cells, MACs and other progenitors has improved cardiovascular outcomes in the acute setting, this is not a viable option in chronic disease. The pharmacological manipulation of MAC/OEC function in vivo however has the potential to significantly improve endothelial repair and thus reduce CVD in this high risk population.

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

    PubMed

    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

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

    PubMed

    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.

  7. 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

  8. Impaired Endothelial Progenitor Cell Mobilization and Dysfunctional Bone Marrow Stroma in Diabetes Mellitus

    PubMed Central

    Rafii, Shahin; Jaspers, Janneke E.; White, Ian A.; Hooper, Andrea T.; Doevendans, Pieter A.; Verhaar, Marianne C.

    2013-01-01

    Background Circulating Endothelial Progenitor Cell (EPC) levels are reduced in diabetes mellitus. This may be a consequence of impaired mobilization of EPC from the bone marrow. We hypothesized that under diabetic conditions, mobilization of EPC from the bone marrow to the circulation is impaired –at least partly– due to dysfunction of the bone marrow stromal compartment. Methods Diabetes was induced in mice by streptozotocin injection. Circulating Sca-1+Flk-1+ EPC were characterized and quantified by flow cytometry at baseline and after mobilization with G-CSF/SCF injections. In vivo hemangiogenic recovery was tested by 5-FU challenge. Interaction within the bone marrow environment between CD34+ hematopoietic progenitor cells (HPC) and supporting stroma was assessed by co-cultures. To study progenitor cell–endothelial cell interaction under normoglycemic and hyperglycemic conditions, a co-culture model using E4Orf1-transfected human endothelial cells was employed. Results In diabetic mice, bone marrow EPC levels were unaffected. However, circulating EPC levels in blood were lower at baseline and mobilization was attenuated. Diabetic mice failed to recover and repopulate from 5-FU injection. In vitro, primary cultured bone marrow stroma from diabetic mice was impaired in its capacity to support human CFU-forming HPC. Finally, hyperglycemia hampered the HPC supportive function of endothelial cells in vitro. Conclusion EPC mobilization is impaired under experimental diabetic conditions and our data suggest that diabetes induces alterations in the progenitor cell supportive capacity of the bone marrow stroma, which could be partially responsible for the attenuated EPC mobilization and reduced EPC levels observed in diabetic patients. PMID:23555959

  9. Nutraceutical augmentation of circulating endothelial progenitor cells and hematopoietic stem cells in human subjects

    PubMed Central

    2010-01-01

    The medical significance of circulating endothelial or hematopoietic progenitors is becoming increasing recognized. While therapeutic augmentation of circulating progenitor cells using G-CSF has resulted in promising preclinical and early clinical data for several degenerative conditions, this approach is limited by cost and inability to perform chronic administration. Stem-Kine is a food supplement that was previously reported to augment circulating EPC in a pilot study. Here we report a trial in 18 healthy volunteers administered Stem-Kine twice daily for a 2 week period. Significant increases in circulating CD133 and CD34 cells were observed at days 1, 2, 7, and 14 subsequent to initiation of administration, which correlated with increased hematopoietic progenitors as detected by the HALO assay. Augmentation of EPC numbers in circulation was detected by KDR-1/CD34 staining and colony forming assays. These data suggest Stem-Kine supplementation may be useful as a stimulator of reparative processes associated with mobilization of hematopoietic and endothelial progenitors. PMID:20377846

  10. Nutraceutical augmentation of circulating endothelial progenitor cells and hematopoietic stem cells in human subjects.

    PubMed

    Mikirova, Nina A; Jackson, James A; Hunninghake, Ron; Kenyon, Julian; Chan, Kyle W H; Swindlehurst, Cathy A; Minev, Boris; Patel, Amit N; Murphy, Michael P; Smith, Leonard; Ramos, Famela; Ichim, Thomas E; Riordan, Neil H

    2010-01-01

    The medical significance of circulating endothelial or hematopoietic progenitors is becoming increasing recognized. While therapeutic augmentation of circulating progenitor cells using G-CSF has resulted in promising preclinical and early clinical data for several degenerative conditions, this approach is limited by cost and inability to perform chronic administration. Stem-Kine is a food supplement that was previously reported to augment circulating EPC in a pilot study. Here we report a trial in 18 healthy volunteers administered Stem-Kine twice daily for a 2 week period. Significant increases in circulating CD133 and CD34 cells were observed at days 1, 2, 7, and 14 subsequent to initiation of administration, which correlated with increased hematopoietic progenitors as detected by the HALO assay. Augmentation of EPC numbers in circulation was detected by KDR-1/CD34 staining and colony forming assays. These data suggest Stem-Kine supplementation may be useful as a stimulator of reparative processes associated with mobilization of hematopoietic and endothelial progenitors. PMID:20377846

  11. Reverse-D-4F Increases the Number of Endothelial Progenitor Cells and Improves Endothelial Progenitor Cell Dysfunctions in High Fat Diet Mice.

    PubMed

    Nana, Yang; Peng, Jiao; Jianlin, Zhang; Xiangjian, Zhang; Shutong, Yao; Enxin, Zhan; Bin, Li; Chuanlong, Zong; Hua, Tian; Yanhong, Si; Yunsai, Du; Shucun, Qin; Hui, Wang

    2015-01-01

    Although high density lipoprotein (HDL) improves the functions of endothelial progenitor cells (EPCs), the effect of HDL ApoAI mimetic peptide reverse-D-4F (Rev-D4F) on EPC mobilization and repair of EPC dysfunctions remains to be studied. In this study, we investigated the effects of Rev-D4F on peripheral blood cell subpopulations in C57 mice treated with a high fat diet and the mechanism of Rev-D4F in improving the function of EPCs impaired by tumor necrosis factor-α (TNF-α). The high fat diet significantly decreased the number of EPCs, EPC migratory functions, and the percentage of lymphocytes in the white blood cells. However, it significantly increased the number of white blood cells, the percentage of monocytes in the white blood cells, and the level of vascular endothelial growth factor (VEGF) and TNF-α in the plasma. Rev-D4F clearly inhibited the effect of the high fat diet on the quantification of peripheral blood cell subpopulations and cytokine levels, and increased stromal cell derived factor 1α (SDF-1α) in the plasma. We provided in vitro evidence that TNF-α impaired EPC proliferation, migration, and tube formation through inactive AKT and eNOS, which was restored by Rev-D4F treatment. In contrast, both the PI3-kinase (PI3K) inhibitor (LY294002) and AKT inhibitor (perifosine) obviously inhibited the restoration of Rev-4F on EPCs impaired by TNF-α. Our results suggested that Rev-D4F increases the quantity of endothelial progenitor cells through increasing the SDF-1α levels and decreasing the TNF-α level of peripheral blood in high fat diet-induced C57BL/6J mice, and restores TNF-α induced dysfunctions of EPCs partly through stimulating the PI3K/AKT signal pathway.

  12. [Promotion of Function of Endothelial Progenitor Cells with Shexiang Baoxin Pill Treatment under Shear Stress].

    PubMed

    Li, Gang; Chen, Yang; Wu, Jiang

    2015-08-01

    The aim of this study was to investigate whether shear stress could promote function of endothelial progenitor cells (EPCs) with Shexiang Baoxin Pill (SBP) treatment in vitro, and to study whether shear stress contributed to vascular injury repair by EPCs. EPCs were isolated and characterized; EPCs' proliferation, migration, adhesion, tube formation and eNOS protein level in vitro were investigated by culturing confluent EPCs in 4 mg/mL SBP under physiological shear stress (15 dyne/cm2) for up to 24 hours. Afterwards, EPCs were transfused into rats after wire-induced carotid artery injury augmented re-endothelialization. The results showed that, compared to the SBP group, the shear stress+SBP group obviously enhanced EPCs proliferation, migration, adhesion, tube formation and eNOS protein expression in vitro (P<0.01). After one week, immunofluorescence staining showed that endothelial regeneration rate obviously enhanced in shear stress+SBP group (P<0.01). The present study demonstrates that shear stress can promote function of endothelial progenitor cells treated with SBP, which improves the vascular injury repair potentials of EPCs. PMID:26710458

  13. Existence of Neural Crest-Derived Progenitor Cells in Normal and Fuchs Endothelial Dystrophy Corneal Endothelium.

    PubMed

    Katikireddy, Kishore Reddy; Schmedt, Thore; Price, Marianne O; Price, Francis W; Jurkunas, Ula V

    2016-10-01

    Human corneal endothelial cells are derived from neural crest and because of postmitotic arrest lack competence to repair cell loss from trauma, aging, and degenerative disorders such as Fuchs endothelial corneal dystrophy (FECD). Herein, we identified a rapidly proliferating subpopulation of cells from the corneal endothelium of adult normal and FECD donors that exhibited features of neural crest-derived progenitor (NCDP) cells by showing absence of senescence with passaging, propensity to form spheres, and increased colony forming efficacy compared with the primary cells. The collective expression of stem cell-related genes SOX2, OCT4, LGR5, TP63 (p63), as well as neural crest marker genes PSIP1 (p75(NTR)), PAX3, SOX9, AP2B1 (AP-2β), and NES, generated a phenotypic footprint of endothelial NCDPs. NCDPs displayed multipotency by differentiating into microtubule-associated protein 2, β-III tubulin, and glial fibrillary acidic protein positive neurons and into p75(NTR)-positive human corneal endothelial cells that exhibited transendothelial resistance of functional endothelium. In conclusion, we found that mitotically incompetent ocular tissue cells contain adult NCDPs that exhibit a profile of transcription factors regulating multipotency and neural crest progenitor characteristics. Identification of normal NCDPs in FECD-affected endothelium holds promise for potential autologous cell therapies. PMID:27639969

  14. Parallel-plate flow chamber and continuous flow circuit to evaluate endothelial progenitor cells under laminar flow shear stress.

    PubMed

    Lane, Whitney O; Jantzen, Alexandra E; Carlon, Tim A; Jamiolkowski, Ryan M; Grenet, Justin E; Ley, Melissa M; Haseltine, Justin M; Galinat, Lauren J; Lin, Fu-Hsiung; Allen, Jason D; Truskey, George A; Achneck, Hardean E

    2012-01-01

    The overall goal of this method is to describe a technique to subject adherent cells to laminar flow conditions and evaluate their response to well quantifiable fluid shear stresses. Our flow chamber design and flow circuit (Fig. 1) contains a transparent viewing region that enables testing of cell adhesion and imaging of cell morphology immediately before flow (Fig. 11A, B), at various time points during flow (Fig. 11C), and after flow (Fig. 11D). These experiments are illustrated with human umbilical cord blood-derived endothelial progenitor cells (EPCs) and porcine EPCs. This method is also applicable to other adherent cell types, e.g. smooth muscle cells (SMCs) or fibroblasts. The chamber and all parts of the circuit are easily sterilized with steam autoclaving. In contrast to other chambers, e.g. microfluidic chambers, large numbers of cells (> 1 million depending on cell size) can be recovered after the flow experiment under sterile conditions for cell culture or other experiments, e.g. DNA or RNA extraction, or immunohistochemistry (Fig. 11E), or scanning electron microscopy. The shear stress can be adjusted by varying the flow rate of the perfusate, the fluid viscosity, or the channel height and width. The latter can reduce fluid volume or cell needs while ensuring that one-dimensional flow is maintained. It is not necessary to measure chamber height between experiments, since the chamber height does not depend on the use of gaskets, which greatly increases the ease of multiple experiments. Furthermore, the circuit design easily enables the collection of perfusate samples for analysis and/or quantification of metabolites secreted by cells under fluid shear stress exposure, e.g. nitric oxide (Fig. 12).

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

    PubMed Central

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

    2014-01-01

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

  16. 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

  17. Cord blood-circulating endothelial progenitors for treatment of vascular diseases.

    PubMed

    Lavergne, M; Vanneaux, V; Delmau, C; Gluckman, E; Rodde-Astier, I; Larghero, J; Uzan, G

    2011-04-01

    Adult peripheral blood (PB) endothelial progenitor cells (EPC) are produced in the bone marrow and are able to integrate vascular structures in sites of neoangiogenesis. EPCs thus represent a potential therapeutic tool for ischaemic diseases. However, use of autologous EPCs in cell therapy is limited by their rarity in adult PB. Cord blood (CB) contains more EPCs than PB, and they are functional after expansion. They form primary colonies that give rise to secondary colonies, each yielding more than 10(7) cells after few passages. The number of endothelial cells obtained from one unit of CB is compatible with potential clinical application. EPC colonies can be securely produced, expanded and cryopreserved in close culture devices and endothelial cells produced in these conditions are functional as shown in different in vitro and in vivo assays. As CB EPC-derived endothelial cells would be allogeneic to patients, it would be of interest to prepare them from ready-existing CB banks. We show that not all frozen CB units from a CB bank are able to generate EPC colonies in culture, and when they do so, number of colonies is lower than that obtained with fresh CB units. However, endothelial cells derived from frozen CB have the same phenotypical and functional properties than those derived from fresh CB. This indicates that CB cryopreservation should be improved to preserve integrity of stem cells other than haematopoietic ones. Feasibility of using CB for clinical applications will be validated in porcine models of ischaemia.

  18. Biocompatibility of porcine small intestinal submucosa and rat endothelial progenitor cells in vitro

    PubMed Central

    Rong, Jian-Jie; Sang, Hong-Fei; Qian, Ai-Min; Meng, Qing-You; Zhao, Tie-Jun; Li, Xiao-Qiang

    2015-01-01

    Objective: This study investigated the biocompatibility of the small intestinal submucosa (SIS) and endothelial progenitor cells (EPCs) by co-cultivating EPCs and SIS in vitro and observing EPC growth on the SIS. Methods: The porcine SIS was prepared and bone marrow mononuclear cells (BMMNCs) were isolated from 3 or 4-week old male SD rats. Cellular morphology was observed by light microscopy and scanning electron microscopy (SEM) and viabilities by the MTT assays. Endothelial progenitor cells (EPCs) were phenotyped by immunocytochemistry, immunofluorescence microscopy and flow cytometry. Vascular lumen formation was evaluated by the Matrigel tube formation assays. EPCs were seeded onto the SIS and production of angiogenin-1 and endothelial cell growth factor (VEGF) by EPCs was examined by ELISA and immunoblotting assays. Results: Light microscopy and SEM showed that the mechanically and chemically treated small intestinal submucosa was composed of cell-free extracellular matrix. Immunohistochemistry, and flow cytometry revealed that the EPCs expressed appropriate surface markers including CD34, CD133, and VEGFR-2. Furthermore, the EPCs formed lumen-like structures and the SIS significantly enhanced the growth of EPCs in vitro. Conclusion: SIS has good biocompatibility with EPCs. SIS pre-seeded with EPCs can be potentially applied as an alternative scaffold material in artificial blood vessel prosthesis. PMID:25973012

  19. 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

  20. Human umbilical cord blood-derived mesenchymal stem cells improve functional recovery through thrombospondin1, pantraxin3, and vascular endothelial growth factor in the ischemic rat brain.

    PubMed

    Park, Hyung Woo; Moon, Hyo-Eun; Kim, Hye-Soo R; Paek, Seung Leal; Kim, Yona; Chang, Jong Wook; Yang, Yoon Sun; Kim, KwanWoo; Oh, Wonil; Hwang, Jae Ha; Kim, Jin Wook; Kim, Dong Gyu; Paek, Sun Ha

    2015-12-01

    Cell therapy is a potential therapeutic method for cerebral ischemia, which remains a serious problem. In the search for more effective therapeutic methods, many kinds of stem cells from various tissues have been developed and tested as candidate therapeutic agents. Among them, human umbilical cord blood (hUCB)-derived mesenchymal stem cells (MSCs) are widely used for cell therapy because of their genetic flexibility. To confirm that they are effective and understand how they affect ischemic neural cells, hUCB-MSCs were directly administered ipsilaterally into an ischemic zone induced by middle cerebral artery occlusion (MCAO). We found that the neurobehavioral performance of the hUCB-MSC group was significantly improved compared with that of the vehicle-injected control group. The infarct was also remarkably smaller in the hUCB-MSC group. Additionally, hUCB-MSC transplantation resulted in a greater number of newly generated cells and angiogenic and tissue repair factors and a lower number of inflammatory events in the penumbra zone. To determine why these events occurred, hUCB-MSCs were assayed under hypoxic and normoxic conditions in vitro. The results showed that hUCB-MSCs exhibit higher expression levels of thrombospondin1, pantraxin3, and vascular endothelial growth factor under hypoxic conditions than under normoxic conditions. These results were found to be correlated with our in vivo immunofluorescent staining results. On the basis of these findings, we suggest that hUCB-MSCs may have a beneficial effect on cerebral ischemia, especially through angiogenesis, neurogenesis, and anti-inflammatory effects, and thus could be used as a therapeutic agent to treat neurological disorders such as cerebral ischemia.

  1. The Secretome of Endothelial Progenitor Cells Promotes Brain Endothelial Cell Activity through PI3-Kinase and MAP-Kinase

    PubMed Central

    Di Santo, Stefano; Seiler, Stefanie; Fuchs, Anna-Lena; Staudigl, Jennifer; Widmer, Hans Rudolf

    2014-01-01

    Background Angiogenesis and vascular remodelling are crucial events in tissue repair mechanisms promoted by cell transplantation. Current evidence underscores the importance of the soluble factors secreted by stem cells in tissue regeneration. In the present study we investigated the effects of paracrine factors derived from cultured endothelial progenitor cells (EPC) on rat brain endothelial cell properties and addressed the signaling pathways involved. Methods Endothelial cells derived from rat brain (rBCEC4) were incubated with EPC-derived conditioned medium (EPC-CM). The angiogenic response of rBCEC4 to EPC-CM was assessed as effect on cell number, migration and tubular network formation. In addition, we have compared the outcome of the in vitro experiments with the effects on capillary sprouting from rat aortic rings. The specific PI3K/AKT inhibitor LY294002 and the MEK/ERK inhibitor PD98059 were used to study the involvement of these two signaling pathways in the transduction of the angiogenic effects of EPC-CM. Results Viable cell number, migration and tubule network formation were significantly augmented upon incubation with EPC-CM. Similar findings were observed for aortic ring outgrowth with significantly longer sprouts. The EPC-CM-induced activities were significantly reduced by the blockage of the PI3K/AKT and MEK/ERK signaling pathways. Similarly to the outcome of the rBCEC4 experiments, inhibition of the PI3K/AKT and MEK/ERK pathways significantly interfered with capillary sprouting induced by EPC-CM. Conclusion The present study demonstrates that EPC-derived paracrine factors substantially promote the angiogenic response of brain microvascular endothelial cells. In addition, our findings identified the PI3K/AKT and MEK/ERK pathways to play a central role in mediating these effects. PMID:24755675

  2. Glioma cells enhance endothelial progenitor cell angiogenesis via VEGFR-2, not VEGFR-1.

    PubMed

    Zhang, Junxia; Zhao, Peng; Fu, Zhen; Chen, Xiaolei; Liu, Ning; Lu, Ailin; Li, Rui; Shi, Lei; Pu, Peiyu; Kang, Chunsheng; You, Yongping

    2008-12-01

    Although potential contribution of endothelial progenitor cells (EPCs) to angiogenesis in glioma has been proposed, the molecular mechanisms of EPCs recruitment to vasculature have not been fully elucidated. Here, we show that the supernatant from glioma cells promotes EPCs angiogenesis via VEGFR-2, not VEGFR-1. Moreover, VEGFR-2 siRNA inhibits VEGFR-2 expression in EPCs, tube formation on matrigel and cell migration. MMP-9 activity and expression and the Akt and ERK phosphorylations are decreased by VEGFR-2 siRNA. Thus, these results indicate that glioma cells enhance EPC angiogenesis via VEGFR-2, not VEGFR-1, mediated by the MMP-9, Akt and ERK signal pathways.

  3. 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

  4. Fabrication of endothelial progenitor cell capture surface via DNA aptamer modifying dopamine/polyethyleneimine copolymer film

    NASA Astrophysics Data System (ADS)

    Li, Xin; Deng, Jinchuan; Yuan, Shuheng; Wang, Juan; Luo, Rifang; Chen, Si; Wang, Jin; Huang, Nan

    2016-11-01

    Endothelial progenitor cells (EPCs) are mainly located in bone marrow and circulate, and play a crucial role in repairmen of injury endothelium. One of the most promising strategies of stents designs were considered to make in-situ endothelialization in vivo via EPC-capture biomolecules on a vascular graft to capture EPCs directly from circulatory blood. In this work, an EPC specific aptamer with a 34 bases single strand DNA sequence was conjugated onto the stent surface via dopamine/polyethyleneimine copolymer film as a platform and linker. The assembled density of DNA aptamer could be regulated by controlling dopamine percentage in this copolymer film. X-ray photoelectron spectroscopy (XPS), water contact angle (WCA) and fluorescence test confirmed the successful immobilization of DNA aptamer. To confirm its biofunctionality and cytocompatibility, the capturing cells ability of the aptamer modified surface and the effects on the growth behavior of human umbilical vein endothelial cells (HUVECs), smooth muscle cells (SMCs) were investigated. The aptamer functionalized sample revealed a good EPC-capture ability, and had a cellular friendly feature for both EPC and EC growth, while not stimulated the hyperplasia of SMCs. And, the co-culture experiment of three types of cells confirmed the specificity capturing of EPCs to aptamer modified surface, rather than ECs and SMCs. These data suggested that this aptamer functionalized surface may have a large potentiality for the application of vascular grafts with targeted endothelialization.

  5. 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

  6. Protective effect of Astragalus polysaccharide on endothelial progenitor cells injured by thrombin.

    PubMed

    Zhang, Xinxia; Yao, Kannan; Ren, Lihong; Chen, Ting; Yao, Dingguo

    2016-01-01

    Several studies have demonstrated that Astragalus polysaccharide (APS) has a protective effect on endothelial cells damaged by various factors. To examine the role of APS in the endothelial inflammatory response, rat bone marrow endothelial progenitor cells (EPCs) were isolated by density gradient centrifugation and identified by immunohistochemistry, then we established a model of inflammatory injury induced by thrombin and measured the effects of APS on EPC viability and proliferation by MTT assays. We also assayed the effect APS had on the inflammatory response, by examining the nuclear factor kappa B (NF-κB) signaling pathway, as well as the expression of intercellular adhesion molecule-1 (ICAM-1), vascular endothelial growth factor (VEGF) and its receptors Flt-1 and KDR. Results demonstrated that EPCs were damaged by thrombin, and APS appeared to inhibit this damage. APS suppressed thrombin-induced ICAM-1 expression by blocking NF-κB signaling in rat bone marrow EPCs, and up-regulating expression of VEGF and its receptors. We believed that APS may be used to treat and prevent EPC injury-related diseases.

  7. 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.

  8. 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

  9. 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

  10. 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

  11. 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.

  12. 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

  13. Inflammasome activation of IL-18 results in endothelial progenitor cell dysfunction in systemic lupus erythematosus.

    PubMed

    Kahlenberg, J Michelle; Thacker, Seth G; Berthier, Celine C; Cohen, Clemens D; Kretzler, Matthias; Kaplan, Mariana J

    2011-12-01

    Systemic lupus erythematosus (SLE) is an autoimmune disease with heterogeneous manifestations including severe organ damage and vascular dysfunction leading to premature atherosclerosis. IFN-α has been proposed to have an important role in the development of lupus and lupus-related cardiovascular disease, partly by repression of IL-1 pathways leading to impairments in vascular repair induced by endothelial progenitor cells (EPCs) and circulating angiogenic cells (CACs). Counterintuitively, SLE patients also display transcriptional upregulation of the IL-1β/IL-18 processing machinery, the inflammasome. To understand this dichotomy and its impact on SLE-related cardiovascular disease, we examined cultures of human and murine control or lupus EPC/CACs to determine the role of the inflammasome in endothelial differentiation. We show that caspase-1 inhibition improves dysfunctional SLE EPC/CAC differentiation into mature endothelial cells and blocks IFN-α-mediated repression of this differentiation, implicating inflammasome activation as a crucial downstream pathway leading to aberrant vasculogenesis. Furthermore, serum IL-18 levels are elevated in SLE and correlate with EPC/CAC dysfunction. Exogenous IL-18 inhibits endothelial differentiation in control EPC/CACs and neutralization of IL-18 in SLE EPC/CAC cultures restores their capacity to differentiate into mature endothelial cells, supporting a deleterious effect of IL-18 on vascular repair in vivo. Upregulation of the inflammasome machinery was operational in vivo, as evidenced by gene array analysis of lupus nephritis biopsies. Thus, the effects of IFN-α are complex and contribute to an elevated risk of cardiovascular disease by suppression of IL-1β pathways and by upregulation of the inflammasome machinery and potentiation of IL-18 activation.

  14. 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

  15. Haptoglobin phenotype may alter endothelial progenitor cell cluster formation in cerebral small vessel disease.

    PubMed

    Rouhl, R P W; van Oostenbrugge, R J; Damoiseaux, J G M C; Debrus-Palmans, L L; Theunissen, R O M F I H; Knottnerus, I L H; Staals, J E A; Delanghe, J R; Tervaert, J W Cohen; Lodder, J

    2009-02-01

    Cerebral small vessel disease results in silent ischemic lesions (SIL) among which is leukoaraiosis. In this process, endothelial damage is probably involved. Endothelial progenitor cells (EPC), are involved in endothelial repair. By restoring the damaged endothelium, EPC could mitigate SIL and cerebral small vessel disease. Haptoglobin 1-1, one of three phenotypes of haptoglobin, relates to SIL and may therefore attenuate the endothelial repair by EPC. Our aim was to quantify EPC number and function and to assess haptoglobin phenotype and its effect on EPC function in patients with a high prevalence of SIL: lacunar stroke patients. We assessed EPC In 42 lacunar stroke patients and 18 controls by flow cytometry and culture with fetal calf serum, patient and control serum. We determined haptoglobin phenotype and cultured EPC with the three different haptoglobin phenotypes. We found that EPC cluster counts were lower in patients (96.9 clusters/well +/- 83.4 (mean +/- SD)), especially in those with SIL (85.0 +/- 64.3), than in controls (174.4 +/- 112.2). Cluster formation was inhibited by patient serum, especially by SIL patient serum, but not by control serum. Patients with haptoglobin 1-1 had less clusters in culture, and when haptoglobin 1-1 was added to EPC cultures, cluster numbers were lower than with the other haptoglobin phenotypes. We conclude that lacunar stroke patients, especially those with SIL, have impaired EPC cluster formation, which may point at decreased endothelial repair potential. The haptoglobin 1-1 phenotype is likely a causative factor in this impairment. PMID:19355924

  16. Mobilization of Circulating Endothelial Progenitor Cells Correlates with the Clinical Course of Hantavirus Disease

    PubMed Central

    Grouls, Stephan; Hettwer, David; Rafat, Neysan; Tönshoff, Burkhard; Zeier, Martin

    2014-01-01

    Infections with hemorrhagic fever viruses are characterized by increased permeability leading to capillary leakage. Hantavirus infection is associated with endothelial dysfunction, and the clinical course is related to the degree of vascular injury. Circulating endothelial progenitor cells (cEPCs) play a pivotal role in the repair of the damaged endothelium. Therefore, we analyzed the number of cEPCs and their mobilizing growth factors in patients suffering from hantavirus disease induced by infection with Puumala virus. The numbers of EPCs of 36 hantavirus-infected patients and age- and gender-matched healthy controls were analyzed by flow cytometry. Concentrations of cEPC-mobilizing growth factors in plasma were determined by enzyme-linked immunosorbent assay. Laboratory parameters were correlated with the number of cEPCs. In patients infected with hantavirus, the number of cEPCs was significantly higher than that in healthy controls. Levels of mobilizing cytokines were upregulated in patients, and the mobilization of cEPCs is paralleled with the normalization of clinical parameters. Moreover, higher levels of cEPCs correlated with higher serum albumin levels and platelet concentrations. Our data indicate that cEPCs may play a role in the repair of hantavirus-induced endothelial damage, thereby influencing the clinical course and the severity of symptoms. PMID:24155401

  17. Decreased Number of Circulating Endothelial Progenitor Cells (CD133+/KDR+) in Patients with Psoriatic Arthritis.

    PubMed

    Batycka-Baran, Aleksandra; Paprocka, Maria; Baran, Wojciech; Szepietowski, Jacek C

    2016-08-23

    Cardiovascular diseases are a major cause of mortality in patients with psoriatic arthritis (PsA), but the precise mechanism of increased cardiovascular risk is unknown. Endothelial dysfunction plays a crucial role in the development of atherosclerosis. Circulating endothelial progenitor cells (CEPCs) contribute to endothelial regeneration and their level may be affected by chronic inflammation. The aim of this study was to evaluate the number of CEPCs in patients with PsA (n = 24) compared with controls (n = 26). CEPCs were identified as CD133+/ KDR+ cells in peripheral blood, using flow cytometry. A significantly decreased number of CEPCs was observed in patients with PsA (p < 0.0001). The number of these cells was significantly, inversely correlated with the severity of skin and joint involvement (Psoriasis Area and Severity Index (PASI), DAS28) and the level of C-reactive protein. We hypothesize that the reduced number of CEPCs may indicate and contribute to the increased cardiovascular risk in patients with PsA.

  18. Circulating endothelial and progenitor cells: Evidence from acute and long-term exercise effects

    PubMed Central

    Koutroumpi, Matina; Dimopoulos, Stavros; Psarra, Katherini; Kyprianou, Theodoros; Nanas, Serafim

    2012-01-01

    Circulating bone-marrow-derived cells, named endothelial progenitor cells (EPCs), are capable of maintaining, generating, and replacing terminally differentiated cells within their own specific tissue as a consequence of physiological cell turnover or tissue damage due to injury. Endothelium maintenance and restoration of normal endothelial cell function is guaranteed by a complex physiological procedure in which EPCs play a significant role. Decreased number of peripheral blood EPCs has been associated with endothelial dysfunction and high cardiovascular risk. In this review, we initially report current knowledge with regard to the role of EPCs in healthy subjects and the clinical value of EPCs in different disease populations such as arterial hypertension, obstructive sleep-apnea syndrome, obesity, diabetes mellitus, peripheral arterial disease, coronary artery disease, pulmonary hypertension, and heart failure. Recent studies have introduced the novel concept that physical activity, either performed as a single exercise session or performed as part of an exercise training program, results in a significant increase of circulating EPCs. In the second part of this review we provide preliminary evidence from recent studies investigating the effects of acute and long-term exercise in healthy subjects and athletes as well as in disease populations. PMID:23272272

  19. Hyperglycaemia exerts deleterious effects on late endothelial progenitor cell secretion actions.

    PubMed

    Zhang, Jie; Zhang, Xiaoyun; Li, Hong; Cui, Xiaodong; Guan, Xiumei; Tang, Kexin; Jin, Chengwen; Cheng, Min

    2013-01-01

    Endothelial progenitor cells (EPCs) play a fundamental role in tissue regeneration and vascular repair both by differentiating into endothelial cells and by secretion of vasoactive substances that promote angiogenesis and maintain vascular homeostasis. It has previously been shown that hyperglycaemia impairs early and late EPC functions, such as differentiation, proliferation and adhesion. However, its role in the regulation of the production of vasoactive substances in EPCs, especially in late EPCs, is less well defined. We investigated the effects of hyperglycaemia on the production of vasodilator, fibrinolytic and angiogenic growth factors, and also on the activity of superoxide dismutase (SOD) in late EPCs. For this purpose, late EPCs were incubated with different concentrations of D-glucose (5-40 mmol/L) for 24 hr. Levels of nitric oxide (NO), tissue plasminogen activator (t-PA), plasminogen activator inhibitor-1 (PAI-1), prostaglandin I(2) (PGI(2)), vascular endothelial growth factor (VEGF) and the activity of SOD were measured by enzyme-linked immunosorbent assay (ELISA). Under high glucose stress conditions, late EPCs exhibited lower levels of NO, t-PA, PAI-1, PGI(2) and VEGF compared to control medium (5 mmol/L glucose). Moreover, high glucose was also observed to decrease the activity of SOD in late EPCs. These results suggest that hyperglycaemia-induced impairment of late EPC secretion functions could contribute to the development of vascular disease in diabetes.

  20. 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.

  1. Dual Angiogenic and Neurotrophic Effects of Bone Marrow–Derived Endothelial Progenitor Cells on Diabetic Neuropathy

    PubMed Central

    Jeong, Jin-Ok; Kim, Mee-Ohk; Kim, Hyongbum; Lee, Min-Young; Kim, Sung-Whan; Ii, Masaaki; Lee, Jung-uek; Lee, Jiyoon; Choi, Yong Jin; Cho, Hyun-Jai; Lee, Namho; Silver, Marcy; Wecker, Andrea; Kim, Dong-Wook; Yoon, Young-sup

    2009-01-01

    Background Endothelial progenitor cells (EPCs) are known to promote neovascularization in ischemic diseases. Recent evidence suggested that diabetic neuropathy is causally related to impaired angiogenesis and deficient growth factors. Accordingly, we investigated whether diabetic neuropathy could be reversed by local transplantation of EPCs. Methods and Results We found that motor and sensory nerve conduction velocities, blood flow, and capillary density were reduced in sciatic nerves of streptozotocin-induced diabetic mice but recovered to normal levels after hind-limb injection of bone marrow–derived EPCs. Injected EPCs were preferentially and durably engrafted in the sciatic nerves. A portion of engrafted EPCs were uniquely localized in close proximity to vasa nervorum, and a smaller portion of these EPCs were colocalized with endothelial cells. Multiple angiogenic and neurotrophic factors were significantly increased in the EPC-injected nerves. These dual angiogenic and neurotrophic effects of EPCs were confirmed by higher proliferation of Schwann cells and endothelial cells cultured in EPC-conditioned media. Conclusions We demonstrate for the first time that bone marrow-derived EPCs could reverse various manifestations of diabetic neuropathy. These therapeutic effects were mediated by direct augmentation of neovascularization in peripheral nerves through long-term and preferential engraftment of EPCs in nerves and particularly vasa nervorum and their paracrine effects. These findings suggest that EPC transplantation could represent an innovative therapeutic option for treating diabetic neuropathy. PMID:19171856

  2. 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

  3. Circulating endothelial progenitor cells are not affected by acute systemic inflammation

    PubMed Central

    Tura, Olga; Haeck, Marlieke L. A.; Short, Abigail; Freyer, Elizabeth; Barclay, G. Robin; Newby, David E.; Mills, Nicholas L.

    2010-01-01

    Vascular injury causes acute systemic inflammation and mobilizes endothelial progenitor cells (EPCs) and endothelial cell (EC) colony-forming units (EC-CFUs). Whether such mobilization occurs as part of a nonspecific acute phase response or is a phenomenon specific to vascular injury remains unclear. We aimed to determine the effect of acute systemic inflammation on EPCs and EC-CFU mobilization in the absence of vascular injury. Salmonella typhus vaccination was used as a model of acute systemic inflammation. In a double-blind randomized crossover study, 12 healthy volunteers received S. typhus vaccination or placebo. Phenotypic EPC populations enumerated by flow cytometry [CD34+VEGF receptor (VEGF)R-2+CD133+, CD14+VEGFR-2+Tie2+, CD45−CD34+, as a surrogate for late outgrowth EPCs, and CD34+CXCR-4+], EC-CFUs, and serum cytokine concentrations (high sensitivity C-reactive protein, IL-6, and stromal-derived factor-1) were quantified during the first 7 days. Vaccination increased circulating leukocyte (9.8 ± 0.6 vs. 5.1 ± 0.2 × 109 cells/l, P < 0.0001), serum IL-6 [0.95 (0–1.7) vs. 0 (0–0) ng/l, P = 0.016], and VEGF-A [60 (45–94) vs. 43 (21–64) pg/l, P = 0.006] concentrations at 6 h and serum high sensitivity C-reactive protein at 24 h [2.7 (1.4–3.6) vs. 0.4 (0.2–0.8) mg/l, P = 0.037]. Vaccination caused a 56.7 ± 7.6% increase in CD14+ cells at 6 h (P < 0.001) and a 22.4 ± 6.9% increase in CD34+ cells at 7 days (P = 0.04). EC-CFUs, putative vascular progenitors, and the serum stromal-derived factor-1 concentration were unaffected throughout the study period (P > 0.05 for all). In conclusion, acute systemic inflammation causes nonspecific mobilization of hematopoietic progenitor cells, although it does not selectively mobilize putative vascular progenitors. We suggest that systemic inflammation is not the primary stimulus for EPC mobilization after acute vascular injury. PMID:20382859

  4. Icariin promotes angiogenic differentiation and prevents oxidative stress-induced autophagy in endothelial progenitor cells.

    PubMed

    Tang, Yubo; Jacobi, Angela; Vater, Corina; Zou, Lijin; Zou, Xuenong; Stiehler, Maik

    2015-06-01

    Reduced tissue levels of endothelial progenitor cells (EPCs) and functional impairment of endothelium are frequently observed in patients with diabetes and cardiovascular disease. The vascular endothelium is specifically sensitive to oxidative stress, and this is one of the mechanisms that causes widespread endothelial dysfunction in most cardiovascular diseases and disorders. Hence attention has increasingly been paid to enhance mobilization and differentiation of EPCs for therapeutic purposes. The aim of this study was to investigate whether Icariin, a natural bioactive component known from traditional Chinese Medicine, can induce angiogenic differentiation and inhibit oxidative stress-induced cell dysfunction in bone marrow-derived EPCs (BM-EPCs), and, if so, through what mechanisms. We observed that treatment of BM-EPCs with Icariin significantly promoted cell migration and capillary tube formation, substantially abrogated hydrogen peroxide (H2 O2 )-induced apoptotic and autophagic programmed cell death that was linked to the reduced intracellular reactive oxygen species levels and restored mitochondrial membrane potential. Icariin downregulated endothelial nitric oxide synthase 3, as well as nicotinamide-adenine dinucleotide phosphate-oxidase expression upon H2 O2 induction. These antiapoptotic and antiautophagic effects of Icariin are possibly mediated by restoring the loss of mammalian target of rapamycin /p70S6K/4EBP1 phosphorylation as well as attenuation of ATF2 and ERK1/2 protein levels after H2 O2 treatment. In summary, favorable modulation of the angiogenesis and redox states in BM-EPCs make Icariin a promising proangiogenic agent both enhancing vasculogenesis and protecting against endothelial dysfunction.

  5. 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

  6. Possible involvement of tumor-producing VEGF-A in the recruitment of lymphatic endothelial progenitor cells from bone marrow.

    PubMed

    Tawada, Masahiro; Hayashi, Shin-Ichiro; Ikegame, Yuka; Nakashima, Shigeru; Yoshida, Kazuhiro

    2014-12-01

    Lymphatic metastasis of human malignant adenocarcinomas is a critical determinant of prognosis. Lymphangiogenesis, the growth of lymphatic vessels, is closely involved in lymphatic metastasis. However, the mechanisms of tumor lymphangiogenesis are not clearly understood. In a previous study, we showed that human gastric cancer MKN45 cells organize neighboring lymphatic vessels via recruitment of bone marrow-derived lymphatic endothelial progenitor cells in a nude mouse xenograft model. The present results also indicated that human colorectal cancer LS174T and breast cancer SK-BR-3 cells promoted lymphangiogenesis as well as the recruitment of lymphatic endothelial progenitor cells from bone marrow. Among growth factors, which are reported to be involved in lymphangiogenesis, only vascular endothelial growth factor (VEGF)-A was extensively secreted by these three types of adenocarcinoma cells in culture. The well-characterized lymphangiogenic factors VEGF-C and VEGF-D in the culture medium of these three types of adenocarcinoma cells were below the detectable levels in ELISA assay. Secretion of epidermal growth factor (EGF) and hepatocyte growth factor (HGF) was not detected. In in vitro culture assay, VEGF-A directly induced the differentiation of bone marrow mononuclear cells into LYVE-1-positive lymphatic endothelial lineage cells. These data collectively suggest the possibility that VEGF-A-rich human adenocarcinomas induce tumor lymphangiogenesis via recruitment of lymphangiogenic endothelial progenitor cells from bone marrow. PMID:25242215

  7. 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

  8. Endothelial progenitor cells: characterization, in vitro expansion, and prospects for autologous cell therapy.

    PubMed

    Smadja, D M; Cornet, A; Emmerich, J; Aiach, M; Gaussem, P

    2007-07-01

    Injection of hematopoietic stem cells or endothelial progenitor cells (EPCs) expanded ex vivo has been shown to augment neovascularization in adult patients, but the precise origin and identity of the cell population responsible for these clinical benefits are controversial. The limited quantity of EPCs in the circulation has been the main obstacle to clinical trials. Several authors have therefore attempted to expand these cells ex vivo in order to obtain a homogeneous cell therapy product. One possible means of expanding EPCs ex vivo is to activate the thrombin receptor PAR-1 with the specific peptide SFLLRN. Indeed, PAR-1 activation promotes cell proliferation and C-X-C chemokine receptor type 4 (CXCR4) dependent migration and differentiation, with an overall angiogenic effect. This review summarizes the results and rationale of clinical trials of angiogenic therapy, the nature of EPCs, the different methods of ex vivo expansion, and current methods of quantification. PMID:17370127

  9. 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.

  10. A new strategy of promoting vascularization of skin substitutes by capturing endothelial progenitor cells automatically.

    PubMed

    Ji, Shi-zhao; Xiao, Shi-chu; Luo, Peng-fei; Huang, Guo-feng; Li, Heng-yu; Zhu, Shi-hui; Xia, Zhao-fan

    2011-10-01

    How to promote vascularization of a skin substitute is the key to successful skin transplantation. Current methods are mainly through releasing angiogenesis-related factors (ARF) or seeding angiogenesis-related cells (ARC), but the efficacy of these methods is not satisfactory, because angiogenesis needs participation of multiple factors, extracellular matrix and related cells. The latest research has demonstrated that endothelial progenitor cells (EPCs) originating from bone marrow and existing in peripheral blood are the key element participating in revascularization of adult tissues. They directly participate in both stem cell vasculogenesis of ischemic tissues and local angiogenesis. We therefore hypothesize whether it is possible to construct a new skin substitute and use it to mobilize EPCs in bone marrow to peripheral circulation and capture EPCs automatically as a simple and effective method of promoting vascularization of the skin substitute for the sake of improving its post-transplant survival. PMID:21840131

  11. Decellularization and Recellularization of Rat Livers With Hepatocytes and Endothelial Progenitor Cells.

    PubMed

    Zhou, Pengcheng; Huang, Yan; Guo, Yibing; Wang, Lei; Ling, Changchun; Guo, Qingsong; Wang, Yao; Zhu, Shajun; Fan, Xiangjun; Zhu, Mingyan; Huang, Hua; Lu, Yuhua; Wang, Zhiwei

    2016-03-01

    Whole-organ decellularization has been identified as a promising choice for tissue engineering. The aim of the present study was to engineer intact whole rat liver scaffolds and repopulate them with hepatocytes and endothelial progenitor cells (EPCs) in a bioreactor. Decellularized liver scaffolds were obtained by perfusing Triton X-100 with ammonium hydroxide. The architecture and composition of the original extracellular matrix were preserved, as confirmed by morphologic, histological, and immunolabeling methods. To determine biocompatibility, the scaffold was embedded in the subcutaneous adipose layer of the back of a heterologous animal to observe the infiltration of inflammatory cells. Hepatocytes were reseeded using a parenchymal injection method and cultured by continuous perfusion. EPCs were reseeded using a portal vein infusion method. Morphologic and functional examination showed that the hepatocytes and EPCs grew well in the scaffold. The present study describes an effective method of decellularization and recellularization of rat livers, providing the foundation for liver engineering and the development of bioartificial livers.

  12. 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.

  13. Endothelial reconstitution by CD34+ progenitors derived from baboon embryonic stem cells.

    PubMed

    Shi, Qiang; Schatten, Gerald; Hodara, Vida; Simerly, Calvin; VandeBerg, John L

    2013-02-01

    In this study, we used a large non-human primate model, the baboon, to establish a step-wise protocol to generate CD34+ endothelial progenitor cells (EPCs) from embryonic stem cells (ESCs) and to demonstrate their reparative effects. Baboon ESCs were sequentially differentiated from embryoid body cultures for 9 days and then were specified into EPCs by culturing them in monolayer for 12 days. The resulting EPCs expressed CD34, CXCR4 and UEA-1, but neither CD31 nor CD117. The EPCs were able to form intact lumen structures when seeded on Matrigel, took up Dil-LDL, and responded to TNF-α. Angioblasts specified in EGM-2 medium and ECGS medium had 6.41 ± 1.16% (n = 3) and 9.32 ± 3.73% CD34+ cells (n = 3). The efficiency of generating CD34+ EPCs did not differ significantly from ECGS to EGM-2 culture media, however, angioblasts specified in ECGS medium expressed a higher percentage of CD34+/CXCR4+ cells (3.49 ± 1.32%, n = 3) than those specified in EGM-2 medium (0.49 ± 0.52%, n = 3). To observe their reparative capacity, we purified CD34+ progenitors after specification by EGM-2 medium; inoculated fluorescently labelled CD34+ EPCs into an arterial segment denuded of endothelium in an ex vivo system. After 14 days of ex vivo culture, the grafted cells had attached and integrated to the denuded surface; in addition, they had matured further and expressed terminally differentiated endothelial markers including CD31 and CD146. In conclusion, we have proved that specified CD34+ EPCs are promising therapeutic agents for repairing damaged vasculature.

  14. 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

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

    PubMed Central

    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 (1H) and high charge and energy (HZE) nuclei (e.g., iron-56Fe) 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 1H- and 56Fe-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 1H- and 56Fe-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

  16. 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

  17. Comparison of Endothelial Cell Phenotypic Markers of Late-Outgrowth Endothelial Progenitor Cells Isolated from Patients with Coronary Artery Disease and Healthy Volunteers

    PubMed Central

    Stroncek, John D.; Grant, Bryan S.; Brown, Melissa A.; Povsic, Thomas J.; Truskey, George A.

    2009-01-01

    The lack of easily isolated autologous endothelial cell (EC) sources is one of the major challenges with vascular tissue engineering interventions. This article examines the isolation and expansion of late-outgrowth endothelial progenitor cells (EPCs) from 50-mL samples of peripheral blood drawn from patients with significant coronary artery disease (CAD) and healthy young adult volunteers. In cases in which late-outgrowth EPCs were successfully isolated, the cells were assayed in vitro for their expression of EC markers, proliferation potential and ability to endothelialize synthetic materials, form new blood vessels, and produce nitric oxide. Late-outgrowth EPCs from patients with CAD and healthy volunteers exhibited critical EC markers and morphological characteristics that were analogous to a control population of human aortic ECs. To our knowledge, this is the first study to examine the suitability of late-outgrowth EPCs from patients with CAD for autologous endothelialization applications. PMID:19435420

  18. 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

  19. Characteristics of bone marrow-derived endothelial progenitor cells in aged mice

    SciTech Connect

    Zhang Wei; Zhang Guoping; Jin Huiming . E-mail: hmjin@shmu.edu.cn; Hu Renming

    2006-09-29

    Evidence for dysfunction of endothelial repair in aged mice was sought by studying the pattern of induced differentiation, quantity, and function of bone marrow-derived endothelial progenitor cells (EPCs) in aged mice. The CD117-positive stem cell population was separated from bone marrow by magnetic activated cell-sorting system (MACS), and EPCs were defined by demonstrating the expression of CD117{sup +}CD34{sup +}Flk-1{sup +} by flow cytometry. After 7 days of culture, the number of clones formed was counted, and proliferation and migration of EPCs were analyzed by MTT[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide] assay and modified Boyden chamber assay. The results demonstrated that compared to the control group, the quantity of bone marrow-derived CD117{sup +} stem cells and EPCs, as well as the proliferation, migration, the number of clones formed, and phagocytotic function of EPCs were significantly reduced in aged mice. There were no significant differences in the morphology and induced differentiation pattern of EPCs between the aged mouse group and the control group. Authors suggest that the dysfunction of EPCs may serve as a surrogate parameter of vascular function in old mice.

  20. Role of vascular endothelial progenitor cells in construction of new vascular loop.

    PubMed

    Zhan, Kuihua; Bai, Lun; Xu, Jianmei

    2013-11-01

    Since bone marrow-derived endothelial progenitor cells (EPCs) have been detected in adult peripheral blood, the mode of vasculogenesis in the adult tissue has caught attention in field of vascularization research. To confirm the role of EPCs in construction of new vascular loop, we took the biomaterial scaffold implanted into adult rat as an experimental model to observe and examine the actions of the EPCs in neovascularization of the material by immunohistochemistry and transmission electron microscopy. Additionally, by establishing a chemotactic migration model for vascular endothelial cells (ECs) and EPCs, the migrations of ECs and EPCs were explored in simulations. The results of 20,000 simulations showed that the number of the vascular loops assisted by the EPCs was 2-5 times that of the vascular sprouts being naturally joined. Based on the results of experiments and simulations, we conclude that the EPCs are able to assist the angiogenic sprouts in joining under the condition of plenty of the EPCs being mobilized, which aggregate at sites close to sprout tips, forming a cell cord and differentiating to ECs in situ, and become vessel segments between neighboring sprouts. This suggests that there is a difference between the adult and embryo in the manner of vasculogenesis and that a small number of EPCs can play an important role to make the new blood vessels achieve rapid functionalization.

  1. Immortalized functional endothelial progenitor cell lines from umbilical cord blood for vascular tissue engineering.

    PubMed

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

    2012-11-01

    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.

  2. Effects of exercise training on endothelial progenitor cells in cardiovascular disease: a systematic review.

    PubMed

    Ribeiro, Fernando; Ribeiro, Ilda P; Alves, Alberto J; do Céu Monteiro, Maria; Oliveira, Nórton L; Oliveira, José; Amado, Francisco; Remião, Fernando; Duarte, José A

    2013-11-01

    This review aimed to examine the effects of exercise training on mobilization of endothelial progenitor cells (EPCs) in patients with cardiovascular disease and to discuss the possible mechanisms involved in the process. A computer-aided search on PubMed and PEDro was conducted to identify relevant studies published up to June 2012. Two reviewers independently selected studies for inclusion and extracted data, namely, quantitative assessment of circulating EPCs. Of the 88 identified studies, 13 met the inclusion criteria. The 13 studies enrolled 648 participants, including patients with chronic heart failure, peripheral artery disease, and coronary artery disease. The exercise characteristics varied largely across the studies: exercise duration ranged from 2 wks to 6 mos, session duration ranged from 20 to 60 mins, and exercise intensity was usually calculated using the maximal heart rate (ranging from 75% to 85%) or the peak/maximum oxygen consumption (60%-70%). All studies used aerobic exercise. The great majority of the 13 studies reported significant effects of different exercise regimens on the number of circulating EPCs. In summary, exercise training seems to increase the number of circulating EPCs, which could contribute to vascular regeneration and angiogenesis. These positive effects of chronic exercise seem to be closely related to the bioavailability of nitric oxide, including increased activity of endothelial nitric oxide synthase and antioxidant enzymes, and activation of matrix metalloproteinase 9.

  3. Specific role of impaired glucose metabolism and diabetes mellitus in endothelial progenitor cell characteristics and function.

    PubMed

    Yiu, Kai-Hang; Tse, Hung-Fat

    2014-06-01

    The disease burden of diabetes mellitus (DM) and its associated cardiovascular complications represent a growing and major global health problem. Recent studies suggest that circulating exogenous endothelial progenitor cells (EPCs) play an important role in endothelial repair and neovascularization at sites of injury or ischemia. Both experimental and clinical studies have demonstrated that hyperglycemia related to DM can induce alterations to EPCs. The reduction and dysfunction of EPCs related to DM correlate with the occurrence and severity of microvascular and macrovascular complications, suggesting a close mechanistic link between EPC dysfunction and impaired vascular function/repair in DM. These alterations to EPCs, likely mediated by multiple pathophysiological mechanisms, including inflammation, oxidative stress, and alterations in Akt and the nitric oxide pathway, affect EPCs at multiple stages: differentiation and mobilization in the bone marrow, trafficking and survival in the circulation, and homing and neovascularization. Several different therapeutic approaches have consequently been proposed to reverse the reduction and dysfunction of EPCs in DM and may represent a novel therapeutic approach to prevent and treat DM-related cardiovascular complications.

  4. 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

  5. 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

  6. Effects of ezetimibe on endothelial progenitor cells and microparticles in high-risk patients.

    PubMed

    Lins, Lívia Campos Amaral; França, Carolina Nunes; Fonseca, Francisco Antonio Helfenstein; Barbosa, Simone Pinto Melo; Matos, Lívia Nascimento; Aguirre, Ana Carolina; Bianco, Henrique Tria; do Amaral, Jonatas Bussador; Izar, Maria Cristina

    2014-09-01

    Imbalance on endothelial turnover can predict cardiovascular outcomes. We aimed at evaluating the effects of lipid-modifying therapies on circulating endothelial progenitor cells (EPCs), endothelial microparticles (EMPs), and platelet microparticles (PMPs) in high cardiovascular risk subjects with elevated C-reactive protein (CRP). Sixty-three individuals with coronary heart disease (CHD) or CHD risk equivalent on stable statin therapy, with LDL-cholesterol <100 mg/dL and CRP ≥ 2.0 mg/L were selected. After a 4-week run-in period with atorvastatin 10 mg, those with persistent CRP ≥ 2.0 mg/L were randomized to another 4-week treatment period with atorvastatin 40 mg, ezetimibe 10 mg or atorvastatin 40 mg/ezetimibe 10 mg. EPC (CD34(+)/CD133(+)/KDR(+)), EMP (CD51(+)), and PMP (CD42(+)/CD31(+)) were quantified by flow cytometry. Atorvastatin 40 mg and atorvastatin 40 mg/ezetimibe 10 mg reduced LDL-cholesterol (P < 0.001, paired T test, vs. baseline). Combined therapy, but not ezetimibe reduced CRP. CD34(+)/KDR(+) EPC were reduced after ezetimibe alone (P = 0.011 vs. baseline, Wilcoxon test) or combined with atorvastatin (P = 0.016 vs. baseline, Wilcoxon test). In addition, ezetimibe increased CD51(+) EMP (P = 0.017 vs. baseline, Wilcoxon test). No correlations between these markers and LDL-cholesterol or CRP were observed. These results contribute to understand the link between inflammation and vascular homeostasis and highlight the broader benefit of statins decreasing inflammation and preventing microparticles release, an effect not observed with ezetimibe alone.

  7. Endothelial Progenitor Cells Predict Cardiovascular Events after Atherothrombotic Stroke and Acute Myocardial Infarction. A PROCELL Substudy

    PubMed Central

    Cuadrado-Godia, Elisa; Regueiro, Ander; Núñez, Julio; Díaz-Ricard, Maribel; Novella, Susana; Oliveras, Anna; Valverde, Miguel A.; Marrugat, Jaume; Ois, Angel; Giralt-Steinhauer, Eva; Sanchís, Juan; Escolar, Ginès; Hermenegildo, Carlos; Roquer, Jaume

    2015-01-01

    Introduction The aim of this study was to determine prognostic factors for the risk of new vascular events during the first 6 months after acute myocardial infarction (AMI) or atherothrombotic stroke (AS). We were interested in the prognostic role of endothelial progenitor cells (EPC) and circulating endothelial cells (CEC) Methods Between February 2009 and July 2012, 100 AMI and 50 AS patients were consecutively studied in three Spanish centres. Patients with previously documented coronary artery disease or ischemic strokes were excluded. Samples were collected within 24h of onset of symptoms. EPC and CEC were studied using flow cytometry and categorized by quartiles. Patients were followed for up to 6 months. NVE was defined as new acute coronary syndrome, transient ischemic attack (TIA), stroke, or any hospitalization or death from cardiovascular causes. The variables included in the analysis included: vascular risk factors, carotid intima-media thickness (IMT), atherosclerotic burden and basal EPC and CEC count. Multivariate survival analysis was performed using Cox regression analysis. Results During follow-up, 19 patients (12.66%) had a new vascular event (5 strokes; 3 TIAs; 4 AMI; 6 hospitalizations; 1 death). Vascular events were associated with age (P = 0.039), carotid IMT≥0.9 (P = 0.044), and EPC count (P = 0.041) in the univariate analysis. Multivariate Cox regression analysis showed an independent association with EPC in the lowest quartile (HR: 10.33, 95%CI (1.22–87.34), P = 0.032] and IMT≥0.9 [HR: 4.12, 95%CI (1.21–13.95), P = 0.023]. Conclusions Basal EPC and IMT≥0.9 can predict future vascular events in patients with AMI and AS, but CEC count does not affect cardiovascular risk. PMID:26332322

  8. Endothelial progenitor cells from peripheral blood support bone regeneration by provoking an angiogenic response.

    PubMed

    Goerke, Sebastian M; Obermeyer, Julia; Plaha, Julia; Stark, G Björn; Finkenzeller, Günter

    2015-03-01

    Neovascularization is crucial for fracture healing and plays an important role in long-time graft survival in tissue engineering applications. Endothelial progenitor cells (EPCs) can be isolated from peripheral blood avoiding donor site morbidity, which makes them attractive for autologous cell-based engineering of neovessels. However, contradictory results are published concerning the vasculogenic potential of this cell type. We could previously show that implanted human endothelial vein cells (HUVECs) gave rise to the formation of a complex functional human neovasculature in a heterotopic (subcutaneous) as well as in an orthotopic (calvarial defect) model of severe combined immunodeficiency (SCID) mice, where vessel formation could even be increased by coimplanting mesenchymal stem cells (MSCs) functioning as perivascular cells. In this study, we investigated whether coimplantation of MSCs which have been predifferentiated in vitro into SMCs (SMC-MSCs) may enable pbEPCs to form blood vessels upon implantation and, if this would be the case, whether the resulting enhanced vascularization may support bone regeneration. For this purpose, pbEPCs and SMC-MSCs were mono- or cocultured in collagen matrices and seeded into scaffolds consisting of decalcified processed bovine cancellous bone (PBCB, Tutobone). Neovascularization and osteogenesis were evaluated using a calvarial bone defect-model in SCID mice. Our experiments could show that the missing vasculogenic potential of pbEPCs is not rescued by coimplantation of SMCs derived from MSCs predifferentiated along the vascular smooth muscle lineage. However, implantation of both cell types alone, or in combination induced an angiogenic response, which correlated in a positive manner with bone formation within the implants.

  9. Bradykinin Preconditioning Improves Therapeutic Potential of Human Endothelial Progenitor Cells in Infarcted Myocardium

    PubMed Central

    Li, Yefei; Yan, Fengdi; Huang, Jie; Ma, Genshan

    2013-01-01

    Objectives Stem cell preconditioning (PC) is a powerful approach in reducing cell death after transplantation. We hypothesized that PC human endothelial progenitor cells (hEPCs) with bradykinin (BK) enhance cell survival, inhibit apoptosis and repair the infarcted myocardium. Methods The hEPCs were preconditioned with or without BK. The hEPCs apoptosis induced by hypoxia along with serum deprivation was determined by annexin V-fluorescein isothiocyanate/ propidium iodide staining. Cleaved caspase-3, Akt and eNOS expressions were determined by Western blots. Caspase-3 activity and vascular endothelial growth factor (VEGF) levels were assessed in hEPCs. For in vivo studies, the survival and cardiomyocytes apoptosis of transplanted hEPCs were assessed using 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindodi- carbocyanine,4-chlorobenzenesul-fonate salt labeled hEPCs and TUNEL staining. Infarct size and cardiac function were measured at 10 days after transplantation, and the survival of transplanted hEPCs were visualized using near-infrared optical imaging. Results In vitro data showed a marked suppression in cell apoptosis following BK PC. The PC reduced caspase-3 activation, increased the Akt, eNOS phosphorylation and VEGF levels. In vivo data in preconditioned group showed a robust cell anti-apoptosis, reduction in infarct size, and significant improvement in cardiac function. The effects of BK PC were abrogated by the B2 receptor antagonist HOE140, the Akt and eNOS antagonists LY294002 and L-NAME, respectively. Conclusions The activation of B2 receptor-dependent PI3K/Akt/eNOS pathway by BK PC promotes VEGF secretion, hEPC survival and inhibits apoptosis, thereby improving cardiac function in vivo. The BK PC hEPC transplantation for stem cell-based therapies is a novel approach that has potential for clinical used. PMID:24312554

  10. 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.

  11. 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

  12. Batroxobin mobilizes circulating endothelial progenitor cells in patients with deep vein thrombosis.

    PubMed

    Lei Zhang; Shi Hong Lu; Li Li; Tao, Yu-Guo; Yong Ling Wan; Senga, Hirobumi; Renchi Yang; Zhong Chao Han

    2011-02-01

    Batroxobin, a thrombin-like enzyme from Bothrops atrox moojeni venom, is associated with the reduction of fibrinogen levels in plasma and the enhancement of anticoagulation and fibrinolysis. In this study, 15 patients with deep vein thrombosis (DVT) achieved successful limb salvage after the administration of batroxobin. We found that the levels of CD34+, CD31+, CD34+/CD31+, and vascular endothelial cadherin (VE-cadherin+) cells had increased in the peripheral blood of patients at 7 days and 14 days after treatment. At 0 day, 7 days, and 14 days, the percentages of CD34+ cells, which are assumed to be hematopoietic stem cells, are 0.39% ± 0.43%, 0.71% ± 0.50%, and 1.11% ± 0.66%, respectively. The levels of CD34+ cells at 14 days are significantly higher than the levels on the first day (P = .004). The levels of CD31+ cells and VE-cadherin+ cells, which represent mature endothelial cells, at 7 days (34.15% ± 11.32%, P = .013; 1.25% ± 1.39%, P = .014) and 14 days (35.21% ± 7.66%, P = .071; 1.85% ± 2.60%, P = .117) were slightly elevated compared with those at 0 day (27.55% ± 8.65%; 0.25 ± 0.39%). The double positive of CD34 and CD31 cells are assumed to be endothelial progenitor cells (EPCs). The levels of CD34+/CD31+ cells at 7 days (0.69% ± 0.50%, P = .001) and 14 days (1.07% ± 0.66%, P = .006) are significantly higher than that on the initial day (0.28% ± 0.30%). The number of CD34+/CD31+ cells significantly increased, indicating that in addition to its role in anticoagulation and fibrinolysis, treatment with batroxobin might simultaneously activate circulating EPCs that might promote the recanalization of the damaged vessel wall. PMID:19825915

  13. Transplanted Endothelial Progenitor Cells Improve Ischemia Muscle Regeneration in Mice by Diffusion Tensor MR Imaging

    PubMed Central

    Bai, Yingying; James, Judy R.; Shlapak, Darya P.

    2016-01-01

    Endothelial progenitor cells (EPCs) play an important role in repairing ischemia tissues. Diffusion tensor imaging (DTI) was applied to detect the architectural organization of skeletal muscle. This study investigated the feasibility and accuracy of using the DTI to evaluate effectiveness of EPCs treatment. Mouse bone marrow-derived EPCs were isolated, cultured, characterized, and transplanted to hindlimb ischemia mice model. DTI was performed on the hindlimb at postischemia time points. The edema regions of diffusion restriction (high signal in diffusion weighted imaging) were decreased in the ischemic muscle of EPCs treated mice after 14 days compared with the controls. These results from DTI show the lower apparent diffusion coefficient and eigenvalues (λ1, λ2, and λ3) and the higher fractional anisotropy and fiber counts of ischemic muscle on 7 and 14 days after EPCs treatment compared to the controls. There was a significant correlation between fiber counts calculated by DTI and survival fibers evaluated by histological section (r = 0.873, P < 0.01). Our study demonstrated that the time frame for muscle fiber regeneration after EPCs transplantation was significantly shortened in vivo. DTI could be a useful tool for noninvasive evaluation of muscle tissue damage and repair in animal models and patient with ischemic diseases. PMID:27656214

  14. 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

  15. 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

  16. Transplanted Endothelial Progenitor Cells Improve Ischemia Muscle Regeneration in Mice by Diffusion Tensor MR Imaging

    PubMed Central

    Bai, Yingying; James, Judy R.; Shlapak, Darya P.

    2016-01-01

    Endothelial progenitor cells (EPCs) play an important role in repairing ischemia tissues. Diffusion tensor imaging (DTI) was applied to detect the architectural organization of skeletal muscle. This study investigated the feasibility and accuracy of using the DTI to evaluate effectiveness of EPCs treatment. Mouse bone marrow-derived EPCs were isolated, cultured, characterized, and transplanted to hindlimb ischemia mice model. DTI was performed on the hindlimb at postischemia time points. The edema regions of diffusion restriction (high signal in diffusion weighted imaging) were decreased in the ischemic muscle of EPCs treated mice after 14 days compared with the controls. These results from DTI show the lower apparent diffusion coefficient and eigenvalues (λ1, λ2, and λ3) and the higher fractional anisotropy and fiber counts of ischemic muscle on 7 and 14 days after EPCs treatment compared to the controls. There was a significant correlation between fiber counts calculated by DTI and survival fibers evaluated by histological section (r = 0.873, P < 0.01). Our study demonstrated that the time frame for muscle fiber regeneration after EPCs transplantation was significantly shortened in vivo. DTI could be a useful tool for noninvasive evaluation of muscle tissue damage and repair in animal models and patient with ischemic diseases.

  17. 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

  18. Bioinformatic analysis of endothelial progenitor cells exposed to folic acid in type 1 diabetes mellitus.

    PubMed

    Fang, D N; He, X D; Li, X H; Jia, H; Li, P Y; Lu, Q; Quan, Z; Wang, Q L

    2014-01-01

    We investigated the effects of type 1 diabetes mellitus (T1DM) on endothelial progenitor cells (EPCs) at the molecular level and assessed the therapeutic potential of folic acid (FA) in DM. We downloaded the gene expression profile of the EPCs from T1DM patients before and after treatment with FA and from healthy controls. We identified the differentially expressed genes (DEGs) in the EPCs from T1DM patients before and after a four-week period of FA treatment and compared them with those obtained from the healthy subjects by using limma package in R language. Then, functional annotation of the DEGs was performed using the online tool Database for Annotation, Visualization and Integrated Discovery (DAVID) based on the Kyoto Encyclopedia of Genes and Genomes database. The expression of 696 genes was altered in the EPCs from T1DM patients compared to those from the healthy controls. These genes were mainly involved in the pathways associated with immune response. FA can normalize majority of the altered gene expression profiles of EPCs from T1DM patients to resemble those of healthy subjects, albeit with some side effects. FA can be a potential therapeutic agent for the treatment of T1DM. However, focused efforts are required to ensure that the dose of FA falls within the permissible pharmacological range.

  19. 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

  20. 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

  1. Transplanted Endothelial Progenitor Cells Improve Ischemia Muscle Regeneration in Mice by Diffusion Tensor MR Imaging.

    PubMed

    Peng, Xin-Gui; Bai, Yingying; James, Judy R; Shlapak, Darya P; Ju, Shenghong

    2016-01-01

    Endothelial progenitor cells (EPCs) play an important role in repairing ischemia tissues. Diffusion tensor imaging (DTI) was applied to detect the architectural organization of skeletal muscle. This study investigated the feasibility and accuracy of using the DTI to evaluate effectiveness of EPCs treatment. Mouse bone marrow-derived EPCs were isolated, cultured, characterized, and transplanted to hindlimb ischemia mice model. DTI was performed on the hindlimb at postischemia time points. The edema regions of diffusion restriction (high signal in diffusion weighted imaging) were decreased in the ischemic muscle of EPCs treated mice after 14 days compared with the controls. These results from DTI show the lower apparent diffusion coefficient and eigenvalues (λ1, λ2, and λ3) and the higher fractional anisotropy and fiber counts of ischemic muscle on 7 and 14 days after EPCs treatment compared to the controls. There was a significant correlation between fiber counts calculated by DTI and survival fibers evaluated by histological section (r = 0.873, P < 0.01). Our study demonstrated that the time frame for muscle fiber regeneration after EPCs transplantation was significantly shortened in vivo. DTI could be a useful tool for noninvasive evaluation of muscle tissue damage and repair in animal models and patient with ischemic diseases. PMID:27656214

  2. [Characterization of endothelial progenitor cells and putative strategies to improve their expansion].

    PubMed

    Smadja, David M; Gaussem, Pascale

    2009-01-01

    Injection of endothelial progenitor cells (EPC) expanded ex vivo has been shown to increase neovascularization in preclinical models of ischemia and in adult patients, but the precise origin and identity of the cell population responsible for these clinical benefits are controversial. Given the potential usefulness of EPC as a cell therapy product, their thorough characterization is of major importance. This review describes the two cell populations currently called EPC and the means to find differential phenotypic markers. We have shown that BMP2/4 are specific markers of late EPC and play a key role in EPC commitment and outgrowth during neovascularization. Several authors have attempted to expand EPC ex vivo in order to obtain a homogeneous cell therapy product. One possible mean of expanding EPC ex vivo is to activate the thrombin receptor PAR-1 with the specific peptide SFLLRN. Indeed, PAR-1 activation increases angiogenic properties of EPC through activation of SDF-1, angiopoietin and IL-8 pathways. This review summarizes the characterization of EPC and different methods of ex vivo expansion. PMID:19527634

  3. Cyclosporine increases ischemia-induced endothelial progenitor cell mobilization through manipulation of the CD26 system.

    PubMed

    Wang, Chao-Hung; Cherng, Wen-Jin; Yang, Ning-I; Hsu, Chia-Ming; Yeh, Chi-Hsiao; Lan, Yii-Jenq; Wang, Jong-Shyan; Verma, Subodh

    2008-03-01

    Cyclosporin A (CsA) improves the success rate of transplantation. The CD26/dipeptidylpeptidase IV (DPP IV) system plays a critical role in mobilizing endothelial progenitor cells (EPCs) from bone marrow. This study investigated whether CsA manipulates CD26/DPP IV activity and increases EPC mobilization. C57BL/6 mice were divided into control and CsA-treated groups. Before and after hindlimb ischemia was induced, circulating EPC number and serum levels of different cytokines were measured. Compared with the controls, CsA treatment significantly increased the blood levels of stroma-derived factor-1alpha and stem cell factor after ischemic stress (P < 0.001). The CsA group displayed a significant increase in the number of circulating EPCs (sca-1+KDR+ and c-kit+CD31+ EPCs, both P < 0.05). In vivo, CsA caused a significant increase in the numbers of EPCs incorporated into the Matrigel and ischemic limbs (P < 0.05). In the peripheral blood, CsA significantly decreased CD26+ cell numbers and attenuated the plasma CD26/DPP IV activity (P < 0.001). Furthermore, short-term CsA treatment significantly improved the perfusion of ischemic limbs and decreased the spontaneous digital amputation rate. In summary, CsA manipulates the mobilization of EPCs into the circulation via the CD26/DPP IV system. Short-term CsA treatment has beneficial effects on angiogenesis of ischemic tissues.

  4. 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

  5. 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

  6. 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

  7. 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

  8. 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

  9. 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

  10. Stromal cell-derived factor-1α prevents endothelial progenitor cells senescence and enhances re-endothelialization of injured arteries via human telomerase reverse transcriptase.

    PubMed

    Shen, Xiaohua; Zhou, Yucheng; Bi, Xukun; Zhang, Jiefang; Fu, Guosheng; Zheng, Hao

    2015-08-01

    Recent studies have suggested that endothelial progenitor subpopulation (EPCs) number and activity were associated with EPCs senescence. Our previous study had shown that stromal cell-derived factor-1alpha (SDF-1α) could prevent EPCs senescence, which may be via telomerase. In this study, we further investigated the role of human telomerase reverse transcriptase (h-TERT) on the protective effect of SDF-1α against senescence. Knockdown h-TERT abrogated the protective effect of SDF-1α and abolished the effects of SDF-1α on migration and proliferation. Moreover, it inhibited EPCs recruitment. In conclusion, h-TERT served a critical role in the progress that SDF-1α prevented EPCs senescence and enhanced re-endothelialization of the injured arteries.

  11. 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

  12. Basic fibroblast growth factor induces VEGF expression in chondrosarcoma cells and subsequently promotes endothelial progenitor cell-primed angiogenesis.

    PubMed

    Tzeng, Huey-En; Chen, Po-Chun; Lin, Kai-Wei; Lin, Chih-Yang; Tsai, Chun-Hao; Han, Shao-Min; Teng, Chieh-Lin; Hwang, Wen-Li; Wang, Shih-Wei; Tang, Chih-Hsin

    2015-07-01

    Chondrosarcoma, a common malignant tumour, develops in bone. Effective adjuvant therapy remains inadequate for treatment, meaning poor prognosis. It is imperative to explore novel remedies. Angiogenesis is a rate-limiting step in progression that explains neovessel formation for blood supply in the tumour microenvironment. Numerous studies indicate that EPCs (endothelial progenitor cells) promote angiogenesis and contribute to tumour growth. bFGF (basic fibroblast growth factor), a secreted cytokine, regulates biological activity, including angiogenesis, and correlates with tumorigenesis. However, the role of bFGF in angiogenesis-related tumour progression by recruiting EPCs in human chondrosarcoma is rarely discussed. In the present study, we found that bFGF induced VEGF (vascular endothelial growth factor) expression via the FGFR1 (fibroblast growth factor receptor 1)/c-Src/p38/NF-κB (nuclear factor κB) signalling pathway in chondrosarcoma cells, thereby triggering angiogenesis of endothelial progenitor cells. Our in vivo data revealed that tumour-secreted bFGF promotes angiogenesis in both mouse plug and chick CAM (chorioallantoic membrane) assays. Xenograft mouse model data, due to bFGF-regulated angiogenesis, showed the bFGF regulates angiogenesis-linked tumour growth. Finally, bFGF was highly expressed in chondrosarcoma patients compared with normal cartilage, positively correlating with VEGF expression and tumour stage. The present study reveals a novel therapeutic target for chondrosarcoma progression.

  13. 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

  14. Electrospun poly(L-lactic acid-co-ɛ-caprolactone) fibers loaded with heparin and vascular endothelial growth factor to improve blood compatibility and endothelial progenitor cell proliferation.

    PubMed

    Chen, Xi; Wang, Jing; An, Qingzhu; Li, Dawei; Liu, Peixi; Zhu, Wei; Mo, Xiumei

    2015-04-01

    Emulsion electrospinning is a convenient and promising method for incorporating proteins and drugs into nanofiber scaffolds. The aim of this study was to fabricate a nanofiber scaffold for anticoagulation and rapid endothelialization. For this purpose, we encapsulated heparin and vascular endothelial growth factor (VEGF) into the core of poly(L-lactic acid-co-ɛ-caprolactone) (P(LLA-CL)) core-shell nanofibers via emulsion electrospinning. The fiber morphology, core-shell structure and hydrophilicity of the nanofiber mats were analyzed by scanning electron microscopy, transmission electron microscopy and water contact angle. The blood compatibility was measured by hemolysis and anticoagulation testing. A CCK-8 assay was performed to study the promotion of endothelial progenitor cell (EPC) growth and was complemented by immunofluorescent staining and SEM. Our study demonstrates that heparin and VEGF can be incorporated into P(LLA-CL) nanofibers via emulsion. The released heparin performed well as an anticoagulant, and the released VEGF promoted EPC growth on the fiber scaffolds. These results imply that electrospun P(LLA-CL) nanofibers containing heparin and VEGF have great potential in the development of vascular grafts in cases where antithrombogenicity and accelerated endothelialization are desirable.

  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. 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

  19. 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

  20. [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

  1. The ROCK inhibitor Y-27632 negatively affects the expansion/survival of both fresh and cryopreserved cord blood-derived CD34+ hematopoietic progenitor cells: Y-27632 negatively affects the expansion/survival of CD34+HSPCs.

    PubMed

    Bueno, Clara; Montes, Rosa; Menendez, Pablo

    2010-06-01

    Cord blood (CB) is an unlimited source of hematopoietic stem and progenitor cells (HSPC). The use of cryopreserved CB-derived CD34+ HSPCs is successful in children and usually leads to rapid hematopoietic recovery upon transplantation. However, current methods for ex vivo expansion of HSPCs still result in a loss of multilineage differentiation potential and current freeze-thawing protocols result in significant cell death and loss of CD34+ HSPCs. The major cause for the loss of viability after slow freezing is apoptosis induced directly by cryoinjury. Very recent reports have demonstrated that Y-27632, a selective and robust ROCK inhibitor is a potent inhibitor of the apoptosis and is efficient in enhancing the post-thaw survival and recovery of different human stem cells including human embryos, hESCs, induced pluripotent stem cells and mesenchymal stem cells. Here, we analyzed the effect of such an inhibitor in CB-derived CD34+ HSPCs. CB-derived CD34+ HSPCs were MACS-isolated and treated with or without 10 microM of Y-27632. The effect of Y-27632 on culture homeostasis was determined in both fresh and cryopreserved CB-derived CD34+ HSPCs. Our results indicate that the Y-27632 not only dramatically inhibits cell expansion of both fresh and cryopreserved CD34+ HSPCs but also impairs survival/recovery of CD34+ HSPCs upon thawing regardless whether Y-27632 is added to both the cryopreservation and the expansion media and or just to the expansion culture medium with or without hematopoietic cytokines. This study identifies for the first time a detrimental effect of Y-27632 on the expansion and survival of both fresh and cryopreserved CB-derived CD34+ HSPCs, suggesting that Y-27632 may have a differential impact on distinct lineage/tissue-specific stem cells. Our data suggest different functions of Y-27632 on human stem cells growing in suspension versus those growing attached to either treated tissue culture plastic or extracellular matrix. We discourage any clinical

  2. Diabetes reduces bone marrow and circulating porcine endothelial progenitor cells, an effect ameliorated by atorvastatin and independent of cholesterol.

    PubMed

    Mohler, Emile R; Shi, Yuquan; Moore, Jonni; Bantly, Andrew; Hamamdzic, Damir; Yoder, Mervin; Rader, Daniel J; Putt, Mary; Zhang, Lifeng; Parmacek, Michael; Wilensky, Robert L

    2009-01-01

    Bone marrow derived endothelial progenitor cells (EPCs) are early precursors of mature endothelial cells which replenish aging and damaged endothelial cells. The authors studied a diabetic swine model to determine if induction of DM adversely affects either bone marrow or circulating EPCs and whether a HMG-CoA reductase inhibitor (statin) improves development and recruitment of EPCs in the absence of cholesterol lowering. Streptozotocin was administered to Yorkshire pigs to induce DM. One month after induction, diabetic pigs were treated with atorvastatin (statin, n = 10), ezetimibe (n = 10) or untreated (n = 10) and evaluated for number of bone marrow and circulating EPCs and femoral artery endothelial function. There was no effect of either medication on cholesterol level. One month after induction of DM prior to administration of drugs, the number of bone marrow and circulating EPCs significantly decreased (P < 0.0001) compared to baseline. Three months after DM induction, the mean proportion of circulating EPCs significantly increased in the atorvastatin group, but not in the control or ezetimibe groups. The control group showed progressive reduction in percentage of flow mediated vasodilatation (no dilatation at 3 months) whereas the atorvastatin group and ezetimibe exhibited vasodilatation, 6% and 4% respectively. DM results in significant impairment of bone marrow and circulating EPCs as well as endothelial function. The effect is ameliorated, in part, by atorvastatin independent of its cholesterol lowering effect. These data suggest a model wherein accelerated atherosclerosis seen with DM may, in part, result from reduction in EPCs which may be ameliorated by treatment with a statin.

  3. Ophthalmic use of blood-derived products.

    PubMed

    Nugent, Ryan B; Lee, Graham A

    2015-01-01

    There is a wide spectrum of blood-derived products that have been used in many different medical and surgical specialties with success. Blood-derived products for clinical use can be extracted from autologous or allogeneic specimens of blood, but recombinant products are also commonly used. A number of blood derivatives have been used for a wide range of ocular conditions, from the ocular surface to the retina. With stringent preparation guidelines, the potential risk of transmission of blood-borne diseases is minimized. We review blood-derived products and how they are improving the management of ocular disease.

  4. 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

  5. Therapeutic Benefit of Bone Marrow–Derived Endothelial Progenitor Cell Transplantation after Experimental Aneurysm Embolization with Coil in Rats

    PubMed Central

    Li, Qianyun; Huang, Jun; Chen, Xi; Chen, Xiaoyan; Zhang, Jun; Wang, Yongting; Yang, Guo-Yuan; Zhu, Wei

    2014-01-01

    Aneurysm embolization with coil is now widely used clinically. However, the recurrence of aneurysms after embolization has always plagued neurosurgeons because the endothelial layer of the aneurysm neck loses its integrity after being embolized by coil. Bone marrow–derived endothelial progenitor cells (BM-EPCs) could be incorporated into injured endothelium and differentiate into mature endothelial cells during vascular repairing processes. The aim of our study is to explore the effects of BM-EPCs on aneurysm repairing and remodeling in a rat embolization model of abdominal aortic aneurysm. BM-EPC proliferation, migration and tube formation were not affected by super-paramagnetic iron oxide nanoparticle (SPIO) labeling compared to the controls (p>0.05). The number of SPIO-labeled cells greatly increased in EPC transplanted rats compared to that of phosphate buffered saline treated rats. SPIO-labeled EPC (SPIO-EPC) are mainly located in the aneurysm neck and surrounded by fibrous tissue. A histology study showed that the aneurysm orifice was closed with neointima and the aneurysm was filled with newly formed fibrous tissue. The SPIO-EPC accumulated in the aneurysm neck, which accelerated focal fibrous tissue remodeling, suggesting that BM-EPCs play a crucial role in repairing and remodeling the aneurysm neck orifice. PMID:24587209

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

    PubMed

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

    2007-08-01

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

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

    PubMed

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

    2016-04-20

    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.

  9. 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

  10. 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

  11. 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

  12. Nitric oxide: a key factor behind the dysfunctionality of endothelial progenitor cells in diabetes mellitus type-2.

    PubMed

    Hamed, Saher; Brenner, Benjamin; Roguin, Ariel

    2011-07-01

    Diabetes mellitus type-2 (DM-2) contributes to atherogenesis by inducing endothelial cell injury and dysfunction. Endothelial progenitor cells (EPCs) are essential to blood vessel formation, can differentiate into mature endothelial cells, and promote the repair of damaged endothelium. In DM-2, the circulating EPC count is low and their functionality is impaired. The mechanisms that underlie this reduced count and impaired functionality are poorly understood. Nitric oxide (NO) is a short-lived signalling molecule that is produced by vascular endothelial cells and participates in the maintenance of vascular tone. NO is also known to participate in other physiological processes, such as cell survival, proliferation, and migration. The bioavailability of NO is reduced in EPCs from DM-2 patients. Interestingly, an inverse relationship exists between the reduction in NO bioavailability in EPCs and the patient's plasma glucose and glycated haemoglobin levels. In addition, NO bioavailability in EPCs correlates with plasma oxidized low-density lipoprotein levels in DM-2. Although this reduction in NO bioavailability could be attributed to oxidative stress in DM-2 patients, it also may be due to impairment of one or more members of the protein signalling cascades that are responsible for NO production. The stimulation of NO production or its signalling cascades in EPCs may increase their numbers and improve their function, thus attenuating endothelium damage, independent of the vasodilatory effects of NO. This review summarizes the metabolic alterations that underlie the molecular mechanisms that may be responsible for EPC decrease and dysfunction in DM-2 with emphasis on the involvement of the NO system.

  13. 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

  14. Effects of simvastatin/ezetimibe on microparticles, endothelial progenitor cells and platelet aggregation in subjects with coronary heart disease under antiplatelet therapy

    PubMed Central

    Camargo, L.M.; França, C.N.; Izar, M.C.; Bianco, H.T.; Lins, L.S.; Barbosa, S.P.; Pinheiro, L.F.; Fonseca, F.A.H.

    2014-01-01

    It is not known whether the addition of ezetimibe to statins adds cardiovascular protection beyond the expected changes in lipid levels. Subjects with coronary heart disease were treated with four consecutive 1-week courses of therapy (T) and evaluations. The courses were: T1, 100 mg aspirin alone; T2, 100 mg aspirin and 40 mg simvastatin/10 mg ezetimibe; T3, 40 mg simvastatin/10 mg ezetimibe, and 75 mg clopidogrel (300 mg initial loading dose); T4, 75 mg clopidogrel alone. Platelet aggregation was examined in whole blood. Endothelial microparticles (CD51), platelet microparticles (CD42/CD31), and endothelial progenitor cells (CD34/CD133; CDKDR/CD133, or CD34/KDR) were quantified by flow cytometry. Endothelial function was examined by flow-mediated dilation. Comparisons between therapies revealed differences in lipids (T2 and T3endothelial function (T2>T1 and T4, P=0.001). Decreased platelet aggregation was observed after aspirin (arachidonic acid, T1endothelial and platelet microparticles, or endothelial progenitor cells. Cardiovascular protection following therapy with simvastatin/ezetimibe seems restricted to lipid changes and improvement of endothelial function not affecting the release of microparticles, mobilization of endothelial progenitor cells or decreased platelet aggregation. PMID:24760119

  15. 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

  16. Impaired vascular remodeling after endothelial progenitor cell transplantation in MMP9-deficient mice suffering cortical cerebral ischemia

    PubMed Central

    Morancho, Anna; Ma, Feifei; Barceló, Verónica; Giralt, Dolors; Montaner, Joan; Rosell, Anna

    2015-01-01

    Endothelial progenitor cells (EPCs) are being investigated for advanced therapies, and matrix metalloproteinase 9 (MMP9) has an important role in stroke recovery. Our aim was to determine whether tissue MMP9 influences the EPC-induced angiogenesis after ischemia. Wild-type (WT) and MMP9-deficient mice (MMP9/KO) were subjected to cerebral ischemia and treated with vehicle or outgrowth EPCs. After 3 weeks, we observed an increase in the peri-infarct vessel density in WT animals but not in MMP9/KO mice; no differences were found in the vehicle-treated groups. Our data suggest that tissue MMP9 has a crucial role in EPC-induced vascular remodeling after stroke. PMID:26219597

  17. Nitric Oxide Donor Molsidomine Positively Modulates Myogenic Differentiation of Embryonic Endothelial Progenitors

    PubMed Central

    Tirone, Mario; Conti, Valentina; Manenti, Fabio; Nicolosi, Pier Andrea; D’Orlando, Cristina; Azzoni, Emanuele

    2016-01-01

    Embryonic VE-Cadherin-expressing progenitors (eVE-Cad+), including hemogenic endothelium, have been shown to generate hematopoietic stem cells and a variety of other progenitors, including mesoangioblasts, or MABs. MABs are vessel-associated progenitors with multilineage mesodermal differentiation potential that can physiologically contribute to skeletal muscle development and regeneration, and have been used in an ex vivo cell therapy setting for the treatment of muscular dystrophy. There is currently a therapeutic need for molecules that could improve the efficacy of cell therapy protocols; one such good candidate is nitric oxide. Several studies in animal models of muscle dystrophy have demonstrated that nitric oxide donors provide several beneficial effects, including modulation of the activity of endogenous cell populations involved in muscle repair and the delay of muscle degeneration. Here we used a genetic lineage tracing approach to investigate whether the therapeutic effect of nitric oxide in muscle repair could derive from an improvement in the myogenic differentiation of eVE-Cad+ progenitors during embryogenesis. We show that early in vivo treatment with the nitric oxide donor molsidomine enhances eVE-Cad+ contribution to embryonic and fetal myogenesis, and that this effect could originate from a modulation of the properties of yolk sac hemogenic endothelium. PMID:27760216

  18. 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

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

    PubMed

    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

  20. 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

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

    PubMed

    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

    2016-08-05

    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.

  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. Advanced glycation end products, carotid atherosclerosis, and circulating endothelial progenitor cells in patients with end-stage renal disease.

    PubMed

    Ueno, Hiroki; Koyama, Hidenori; Fukumoto, Shinya; Tanaka, Shinji; Shoji, Takuhito; Shoji, Tetsuo; Emoto, Masanori; Tahara, Hideki; Inaba, Masaaki; Kakiya, Ryusuke; Tabata, Tsutomu; Miyata, Toshio; Nishizawa, Yoshiki

    2011-04-01

    Numbers of endothelial progenitor cells (EPCs) have been shown to be decreased in subjects with end-stage renal disease (ESRD), the mechanism of which remained poorly understood. In this study, mutual association among circulating EPC levels, carotid atherosclerosis, serum pentosidine, and skin autofluorescence, a recently established noninvasive measure of advanced glycation end products accumulation, was examined in 212 ESRD subjects undergoing hemodialysis. Numbers of circulating EPCs were measured as CD34+ CD133+ CD45(low) VEGFR2+ cells and progenitor cells as CD34+ CD133+ CD45(low) fraction by flow cytometry. Skin autofluorescence was assessed by the autofluorescence reader; and serum pentosidine, by enzyme-linked immunosorbent assay. Carotid atherosclerosis was determined as intimal-medial thickness (IMT) measured by ultrasound. Circulating EPCs were significantly and inversely correlated with skin autofluorescence in ESRD subjects (R = -0.216, P = .002), but not with serum pentosidine (R = -0.079, P = .25). Circulating EPCs tended to be inversely associated with IMT (R = -0.125, P = .069). Intimal-medial thickness was also tended to be correlated positively with skin autofluorescence (R = 0.133, P = .054) and significantly with serum pentosidine (R = 0.159, P = .019). Stepwise multiple regression analyses reveal that skin autofluorescence, but not serum pentosidine and IMT, was independently associated with low circulating EPCs. Of note, skin autofluorescence was also inversely and independently associated with circulating progenitor cells. Thus, tissue accumulated, but not circulating, advanced glycation end products may be a determinant of a decrease in circulating EPCs in ESRD subjects.

  4. Advanced glycation end products, carotid atherosclerosis, and circulating endothelial progenitor cells in patients with end-stage renal disease.

    PubMed

    Ueno, Hiroki; Koyama, Hidenori; Fukumoto, Shinya; Tanaka, Shinji; Shoji, Takuhito; Shoji, Tetsuo; Emoto, Masanori; Tahara, Hideki; Inaba, Masaaki; Kakiya, Ryusuke; Tabata, Tsutomu; Miyata, Toshio; Nishizawa, Yoshiki

    2011-04-01

    Numbers of endothelial progenitor cells (EPCs) have been shown to be decreased in subjects with end-stage renal disease (ESRD), the mechanism of which remained poorly understood. In this study, mutual association among circulating EPC levels, carotid atherosclerosis, serum pentosidine, and skin autofluorescence, a recently established noninvasive measure of advanced glycation end products accumulation, was examined in 212 ESRD subjects undergoing hemodialysis. Numbers of circulating EPCs were measured as CD34+ CD133+ CD45(low) VEGFR2+ cells and progenitor cells as CD34+ CD133+ CD45(low) fraction by flow cytometry. Skin autofluorescence was assessed by the autofluorescence reader; and serum pentosidine, by enzyme-linked immunosorbent assay. Carotid atherosclerosis was determined as intimal-medial thickness (IMT) measured by ultrasound. Circulating EPCs were significantly and inversely correlated with skin autofluorescence in ESRD subjects (R = -0.216, P = .002), but not with serum pentosidine (R = -0.079, P = .25). Circulating EPCs tended to be inversely associated with IMT (R = -0.125, P = .069). Intimal-medial thickness was also tended to be correlated positively with skin autofluorescence (R = 0.133, P = .054) and significantly with serum pentosidine (R = 0.159, P = .019). Stepwise multiple regression analyses reveal that skin autofluorescence, but not serum pentosidine and IMT, was independently associated with low circulating EPCs. Of note, skin autofluorescence was also inversely and independently associated with circulating progenitor cells. Thus, tissue accumulated, but not circulating, advanced glycation end products may be a determinant of a decrease in circulating EPCs in ESRD subjects. PMID:20494372

  5. 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

  6. 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.

  7. 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

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

    PubMed

    Duttenhoefer, Fabian; de Freitas, Rafael Lara; Loibl, Markus; Bittermann, Gido; Richards, R Geoff; 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

  9. 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.

  10. 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.

  11. 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

  12. 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

  13. 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

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

    PubMed Central

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

    2016-01-01

    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

  15. GSK3β inhibition activates the CDX/HOX pathway and promotes hemogenic endothelial progenitor differentiation from human pluripotent stem cells.

    PubMed

    Kitajima, Kenji; Nakajima, Marino; Kanokoda, Mai; Kyba, Michael; Dandapat, Abhijit; Tolar, Jakub; Saito, Megumu K; Toyoda, Masashi; Umezawa, Akihiro; Hara, Takahiko

    2016-01-01

    WNT/β-CATENIN signaling promotes the hematopoietic/endothelial differentiation of human embryonic stem cells and human induced pluripotent stem cells (hiPSCs). The transient addition of a GSK3β inhibitor (GSKi) has been found to facilitate in vitro endothelial cell differentiation from hESCs/hiPSCs. Because hematopoietic and endothelial cells are derived from common progenitors (hemogenic endothelial progenitors [HEPs]), we examined the effect of transient GSKi treatment on hematopoietic cell differentiation from hiPSCs. We found that transient GSKi treatment at the start of hiPSC differentiation induction altered the gene expression profile of the cells. Multiple CDX/HOX genes, which are expressed in the posterior mesoderm of developing embryos, were significantly upregulated by GSKi treatment. Further, inclusion of the GSKi in a serum- and stroma-free culture with chemically defined medium efficiently induced HEPs, and the HEPs gave rise to various lineages of hematopoietic and endothelial cells. Therefore, transient WNT/β-CATENIN signaling triggers activation of the CDX/HOX pathway, which in turn confers hemogenic posterior mesoderm identity to differentiating hiPSCs. These data enhance our understanding of human embryonic hematopoietic/endothelial cell development and provide a novel in vitro system for inducing the differentiation of hematopoietic cells from hiPSCs. PMID:26477526

  16. A reversal of age-dependent proliferative capacity of endothelial progenitor cells from different species origin in in vitro condition

    PubMed Central

    Hassanpour, Mehdi; Cheraghi, Omid; Siavashi, Vahid; Rahbarghazi, Reza; Nouri, Mohammad

    2016-01-01

    Introduction: A large number of cardiovascular disorders and abnormalities, notably accelerated vascular deficiencies could be related to aging changes and increased length of life. During the past decades, the discovery of different stem cells facilitates ongoing attempts for attenuating many disorders, especially in vascular beds. Endothelial progenitor cells (EPCs) are a subtype of stem cells that have potent capacity to differentiate into mature endothelial cells (ECs). However, some documented studies reported an age-related decline in proliferation and function of many stem cells. There is no data on aging effect upon proliferation and morphological feature of EPCs. Methods: To show aging effect on EPCs proliferation and multipotentiality, bone marrow samples were provided from old and young cases in three different species; human, mouse and dog. After 7 days of culture, the cell morphology and clonogenic capacity were evaluated. We also calculated the mean number of colonies both in bone marrow samples from old and young subjects. To confirm the cell phenotype, isolated cells were immune-phenotyped by a panel of antibodies against Tie-2, CD133 and CD309 markers. Results: Our results showed that EPCs exhibited prominent spindle form in all bone marrow samples from young cases while the cell shape became more round by aging. Notably, the number of colonies was reduced in aged samples as compared to parallel young subject samples (P < 0.05). We also detected that the expression of endothelial related markers diminished by aging. Conclusion: The results of this study suggest that the age-related vascular abnormalities could be presumably related to the decline in stemness capacity of EPCs. PMID:27777694

  17. Development of serum-free quality and quantity control culture of colony-forming endothelial progenitor cell for vasculogenesis.

    PubMed

    Masuda, Haruchika; Iwasaki, Hiroto; Kawamoto, Atsuhiko; Akimaru, Hiroshi; Ishikawa, Masakazu; Ii, Masaaki; Shizuno, Tomoko; Sato, Atsuko; Ito, Rie; Horii, Miki; Ishida, Hideyuki; Kato, Shunichi; Asahara, Takayuki

    2012-02-01

    Quantitative and qualitative impairment of endothelial progenitor cells (EPCs) limits the efficacy of autologous cell therapy in patients with cardiovascular diseases. Here, we developed a serum-free quality and quantity control culture system for colony-forming EPCs to enhance their regenerative potential. A culture with serum-free medium containing stem cell factor, thrombopoietin, vascular endothelial growth factor, interleukin-6, and Flt-3 ligand was determined as optimal quality and quantity culture (QQc) in terms of the most vasculogenic colony-forming EPC expansion, evaluated by the newly established EPC colony formation assay. The QQc of umbilical cord blood-CD133(+) cells for 7 days produced a 52.9-fold increase in total cell number and 3.28-fold frequency in definitive EPC colony development, resulting in a 203.9-fold increase in estimated total definitive EPC colony number in vitro. Pre- or post-QQc cells were intramyocardially transplanted into nude rats with myocardial infarction (MI). Echocardiographic and micromanometer-tipped conductance catheter examinations 28 days post-MI revealed significant preservation of left ventricular (LV) function in rats receiving pre- or post-QQc cells compared with those receiving phosphate-buffered saline. Assessments of global LV contractility indicated a dose-dependent effect of pre- or post-QQc cells and the superior potency of post-QQc cells over pre-QQc cells. Furthermore, immunohistochemistry showed more abundant formation of both human and rat endothelial cells and cardiomyocytes in the infarcted myocardium following transplantation of post-QQc cells compared with pre-QQc cells. Our optimal serum-free quality and quantity culture may enhance the therapeutic potential of EPCs in both quantitative and qualitative aspects for cardiovascular regeneration.

  18. Rapid synthesis of water-dispersible superparamagnetic iron oxide nanoparticles by a microwave-assisted route for safe labeling of endothelial progenitor cells.

    PubMed

    Carenza, Elisa; Barceló, Verónica; Morancho, Anna; Montaner, Joan; Rosell, Anna; Roig, Anna

    2014-08-01

    We synthesize highly crystalline citrate-coated iron oxide superparamagnetic nanoparticles that are stable and readily dispersible in water by an extremely fast microwave-assisted route and investigate the uptake of magnetic nanoparticles by endothelial cells. Nanoparticles form large aggregates when added to complete endothelial cell medium. The size of the aggregates was controlled by adjusting the ionic strength of the medium. The internalization of nanoparticles into endothelial cells was then investigated by transmission electron microscopy, magnetometry and chemical analysis, together with cell viability assays. Interestingly, a sevenfold more efficient uptake was found for systems with larger nanoparticle aggregates, which also showed significantly higher magnetic resonance imaging effectiveness without compromising cell viability and functionality. We are thus presenting an example of a straightforward microwave synthesis of citrate-coated iron oxide nanoparticles for safe endothelial progenitor cell labeling and good magnetic resonance cell imaging with potential application for magnetic cell guidance and in vivo cell tracking.

  19. Histone Deacetylase Inhibition Enhances Self Renewal and Cardioprotection by Human Cord Blood-Derived CD34+ Cells

    PubMed Central

    Burba, Ilaria; Colombo, Gualtiero I.; Staszewsky, Lidia Irene; De Simone, Marco; Devanna, Paolo; Nanni, Simona; Avitabile, Daniele; Molla, Fabiola; Cosentino, Simona; Russo, Ilaria; De Angelis, Noeleen; Soldo, Annarita; Biondi, Antonella; Gambini, Elisa; Gaetano, Carlo; Farsetti, Antonella; Pompilio, Giulio; Latini, Roberto; Capogrossi, Maurizio C.; Pesce, Maurizio

    2011-01-01

    Background Use of peripheral blood- or bone marrow-derived progenitors for ischemic heart repair is a feasible option to induce neo-vascularization in ischemic tissues. These cells, named Endothelial Progenitors Cells (EPCs), have been extensively characterized phenotypically and functionally. The clinical efficacy of cardiac repair by EPCs cells remains, however, limited, due to cell autonomous defects as a consequence of risk factors. The devise of “enhancement” strategies has been therefore sought to improve repair ability of these cells and increase the clinical benefit. Principal Findings Pharmacologic inhibition of histone deacetylases (HDACs) is known to enhance hematopoietic stem cells engraftment by improvement of self renewal and inhibition of differentiation in the presence of mitogenic stimuli in vitro. In the present study cord blood-derived CD34+ were pre-conditioned with the HDAC inhibitor Valproic Acid. This treatment affected stem cell growth and gene expression, and improved ischemic myocardium protection in an immunodeficient mouse model of myocardial infarction. Conclusions Our results show that HDAC blockade leads to phenotype changes in CD34+ cells with enhanced self renewal and cardioprotection. PMID:21789227

  20. Absence of a relationship between immunophenotypic and colony enumeration analysis of endothelial progenitor cells in clinical haematopoietic cell sources

    PubMed Central

    Tura, Olga; Barclay, G Robin; Roddie, Huw; Davies, John; Turner, Marc L

    2007-01-01

    Background The discovery of adult endothelial progenitor cells (EPC) offers potential for vascular regenerative therapies. The expression of CD34 and VEGFR2 by EPC indicates a close relationship with haematopoietic progenitor cells (HPC), and HPC-rich sources have been used to treat cardiac and limb ischaemias with apparent clinical benefit. However, the laboratory characterisation of the vasculogenic capability of potential or actual therapeutic cell autograft sources is uncertain since the description of EPC remains elusive. Various definitions of EPC based on phenotype and more recently on colony formation (CFU-EPC) have been proposed. Methods We determined EPC as defined by proposed phenotype definitions (flow cytometry) and by CFU-EPC in HPC-rich sources: bone marrow (BM); cord blood (CB); and G-CSF-mobilised peripheral blood (mPB), and in HPC-poor normal peripheral blood (nPB). Results As expected, the highest numbers of cells expressing the HPC markers CD34 or CD133 were found in mPB and least in nPB. The proportions of CD34+ cells co-expressing CD133 is of the order mPB>CB>BM≈nPB. CD34+ cells co-expressing VEGFR2 were also most frequent in mPB. In contrast, CFU-EPC were virtually absent in mPB and were most readily detected in nPB, the source lowest in HPC. Conclusion HPC sources differ in their content of putative EPC. Normal peripheral blood, poor in HPC and in HPC-related phenotypically defined EPC, is the richest source of CFU-EPC, suggesting no direct relationship between the proposed EPC immunophenotypes and CFU-EPC potential. It is not apparent whether either of these EPC measurements, or any, is an appropriate indicator of the therapeutic vasculogenic potential of autologous HSC sources. PMID:17640360

  1. Fetal exposure to a diabetic intrauterine environment resulted in a failure of cord blood endothelial progenitor cell adaptation against chronic hypoxia

    PubMed Central

    Dincer, U Deniz

    2015-01-01

    Gestational diabetes mellitus (GDM) has long-term health consequences, and fetal exposure to a diabetic intrauterine environment increases cardiovascular risk for her adult offspring. Some part of this could be related to their endothelial progenitor cells (EPCs). Understanding the vessel-forming ability of human umbilical cord blood (HUCB)-derived endothelial colony-forming cells (ECFCs) against pathological stress such as GDM response to hypoxia could generate new therapeutic strategies. This study aims to investigate the role of chronic hypoxia in EPCs functional and vessel-forming ability in GDM subjects. Each ECFC was expressed in endothelial and pro-angiogenic specific markers, namely endothelial nitric oxide synthase (eNOS), platelet (PECAM-1) endothelial cell adhesion molecule 1, vascular endothelial-cadherin CdH5 (Ca-dependent cell adhesion molecule), vascular endothelial growth factor A, (VEGFA) and insulin-like growth factor 1 (IGF1). Chronic hypoxia did not affect CdH5, but PECAM1 MRNA expressions were increased in control and GDM subjects. Control hypoxic and GDM normoxic VEGFA MRNA expressions and hypoxia-inducible factor 1-alpha (HIF1α) protein expressions were significantly increased in HUCB ECFCs. GDM resulted in most failure of HUCB ECFC adaptation and eNOS protein expressions against chronic hypoxia. Chronic hypoxia resulted in an overall decline in HUCB ECFCs’ proliferative ability due to reduction of clonogenic capacity and diminished vessel formation. Furthermore, GDM also resulted in most failure of cord blood ECFC adaptation against chronic hypoxic environment. PMID:25565870

  2. GTP cyclohydrolase I prevents diabetic-impaired endothelial progenitor cells and wound healing by suppressing oxidative stress/thrombospondin-1.

    PubMed

    Tie, Lu; Chen, Lu-Yuan; Chen, Dan-Dan; Xie, He-Hui; Channon, Keith M; Chen, Alex F

    2014-05-15

    Endothelial progenitor cell (EPC) dysfunction is a key contributor to diabetic refractory wounds. Endothelial nitric oxide synthase (eNOS), which critically regulates the mobilization and function of EPCs, is uncoupled in diabetes due to decreased cofactor tetrahydrobiopterin (BH4). We tested whether GTP cyclohydrolase I (GTPCH I), the rate-limiting enzyme of BH4 synthesis, preserves EPC function in type 1 diabetic mice. Type 1 diabetes was induced in wild-type (WT) and GTPCH I transgenic (Tg-GCH) mice by intraperitoneal injection of streptozotocin (STZ). EPCs were isolated from the peripheral blood and bone marrow of WT, Tg-GCH, and GTPCH I-deficient hph-1 mice. The number of EPCs was significantly lower in STZ-WT mice and hph-1 mice and was rescued in STZ Tg-GCH mice. Furthermore, GTPCH I overexpression improved impaired diabetic EPC migration and tube formation. EPCs from WT, Tg-GCH, and STZ-Tg-GCH mice were administered to diabetic excisional wounds and accelerated wound healing significantly, with a concomitant augmentation of angiogenesis. Flow cytometry measurements showed that intracellular nitric oxide (NO) levels were reduced significantly in STZ-WT and hph-1 mice, paralleled by increased superoxide anion levels; both were rescued in STZ-Tg-GCH mice. Western blot analysis revealed that thrombospondin-1 (TSP-1) was significantly upregulated in the EPCs of STZ-WT mice and hph-1 mice and suppressed in STZ-treated Tg-GCH mice. Our results demonstrate that the GTPCH I/BH4 pathway is critical to preserve EPC quantity, function, and regenerative capacity during wound healing in type 1 diabetic mice at least partly through the attenuation of superoxide and TSP-1 levels and augmentation of NO level.

  3. 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

  4. 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

  5. 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

  6. 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

  7. Interleukin 8 is differently expressed and modulated by PAR-1 activation in early and late endothelial progenitor cells.

    PubMed

    Smadja, David M; Bièche, Ivan; Susen, Sophie; Mauge, Laetitia; Laurendeau, Ingrid; d'Audigier, Clément; Grelac, Françoise; Emmerich, Joseph; Aiach, Martine; Gaussem, Pascale

    2009-08-01

    The proinflammatory chemokine interleukin 8 exerts potent angiogenic effects on endothelial cells by interacting with its receptors CXCR1 and CXCR2. As thrombin is also a potent inflammatory factor, and as endothelial progenitor cells (EPC) express functional PAR-1 thrombin receptor, we examined whether PAR-1 stimulation interferes with the IL-8 pathway in EPC. EPC were obtained from adult blood (AB) and cord blood (CB). The effect of PAR-1 stimulation by the peptide SFLLRN on IL-8, CXCR1 and CXCR2 expression was examined by RTQ-PCR and at the protein level in AB and CB late EPC and in AB early EPC. Specific siRNA was used to knock down PAR-1 expression. The IL-8 gene was expressed strongly in AB early EPC and moderately in late EPC. In contrast, CXCR1 and CXCR2 gene expression was restricted to AB early EPC. The IL-8 level in AB early EPC conditioned medium was high in basal conditions and did not change after PAR-1 activation. By contrast, IL-8 secretion by late EPC was low in basal conditions and strongly up-regulated upon PAR-1 activation. PAR-1 activation induced a number of genes involved in activating protein-1 (AP-1) and nuclear factor (NF)-kappaB pathways. Conditioned medium of PAR-1-activated late EPC enhanced the migratory potential of early EPC, and this effect was abrogated by blocking IL-8. Target-specific siRNA-induced PAR-1 knockdown, and fully inhibited PAR-1-induced IL-8 synthesis. In conclusion, PAR-1 activation induces IL-8 synthesis by late EPC. This could potentially enhance cooperation between late and early EPC during neovascularization, through a paracrine effect. PMID:18657231

  8. In vitro evaluation of endothelial progenitor cells from adipose tissue as potential angiogenic cell sources for bladder angiogenesis.

    PubMed

    Zhou, Liuhua; Xia, Jiadong; Qiu, Xuefeng; Wang, Pengji; Jia, Ruipeng; Chen, Yun; Yang, Bin; Dai, Yutian

    2015-01-01

    Autologous endothelial progenitor cells (EPCs) might be alternative angiogenic cell sources for vascularization of tissue-engineered bladder, while isolation and culture of EPCs from peripheral blood in adult are usually time-consuming and highly inefficient. Recent evidence has shown that EPCs also exist in the adipose tissue. As adipose tissue is plentiful in the human body and can be easily harvested through a minimally invasive method, the aim of this study was to culture and characterize EPCs from adipose tissue (ADEPCs) and investigate their potential for the neovascularization of tissue-engineered bladder. Adipose stromal vascular fraction (SVF) was isolated and used for the culture of ADEPCs and adipose derived stem cells (ADSCs). After SVF was cultured for one week, ADEPCs with typical cobblestone morphology emerged and could be isolated from ADSCs according to their different responses to trypsinization. Rat bladder smooth muscle cells (RBSMCs) were isolated and cultured from rat bladder. RBSMCs exhibited typical spindle-shaped morphology. ADEPCs had higher proliferative potential than ADSCs and RBSMCs. ADEPCs stained positive for CD34, Stro-1, VEGFR-2, eNOS and CD31 but negative for α-SMA, CD14 and CD45. ADSCs stained positive for CD34, Stro-1 and α-SMA but negative for VEGFR-2, eNOS, CD31, CD14 and CD45. RBSMCs stained only positive for α-SMA. ADEPCs could be expanded from a single cell at an early passage to a cell cluster containing more than 10,000 cells. ADEPCs were able to uptake DiI-Ac-LDL, bind UEA-1 and form capillary-like structures in three-dimensional scaffolds (Matrigel and bladder acellular matrix). ADEPCs were also able to enhance the human umbilical vein endothelial cells' capability of capillary-like tube formation on Matrigel. Additionally, significantly higher levels of mRNA and protein of vascular endothelial growth factor were found in ADEPCs than in RBSMCs. These results suggest the potential use of ADEPCs as angiogenic cell

  9. 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

  10. 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

  11. SDF-1α-induced dual pairs of E-selectin/ligand mediate endothelial progenitor cell homing to critical ischemia

    PubMed Central

    Liu, Zhao-Jun; Tian, Runxia; Li, Yan; Zhang, Leiming; Shao, Hongwei; Yang, Cuixia; Velazquez, Omaida C.

    2016-01-01

    Homing of endothelial progenitor cells (EPC) to the ischemic tissues is a key event in neovascularization and tissue regeneration. In response to ischemic insult, injured tissues secrete several chemo-cytokines, including stromal cell-derived factor-1α (SDF-1α), which triggers mobilization and homing of bone marrow-derived EPC (BMD-EPC). We previously reported that SDF-1α-induced EPC homing is mediated by a panel of adhesion molecules highly or selectively expressed on the activated endothelium in ischemic tissues, including E-selectin. Elevated E-selectin on wound vasculature serve as docking sites for circulating EPC, which express counterpart E-selectin ligands. Here, we show that SDF-1α presented in wound tissue and released into circulation can act both locally and remotely to induce ischemic tissue endothelium and BMD-EPC to express both E-selectin and its ligands. By performing BM transplantation using E-selectin−/− and E-selectin+/+ mice as the donors and recipients respectively, we demonstrate that upregulated dual E-selectin/ligand pairs reciprocally expressed on ischemic tissue endothelium and BMD-EPC act as double-locks to secure targeted EPC- endothelium interactions by which to facilitate EPC homing and promote neovascularization and tissue repair. These findings describe a novel mechanism for BMD-EPC homing and indicate that dual E-selectin/ligand pairs may be effective targets/tools for therapeutic neovascularization and targeted cell delivery. PMID:27713493

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

    PubMed

    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

  13. 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

  14. Gender differences in circulating endothelial progenitor cell colony-forming capacity and migratory activity in middle-aged adults.

    PubMed

    Hoetzer, Greta L; MacEneaney, Owen J; Irmiger, Heather M; Keith, Rebecca; Van Guilder, Gary P; Stauffer, Brian L; DeSouza, Christopher A

    2007-01-01

    Middle-aged women have a lower prevalence and incidence of cardiovascular events compared with men. The mechanisms responsible for this gender-specific difference are unclear. Numeric and functional impairments of bone marrow-derived circulating endothelial progenitor cells (EPCs) are associated with increased cardiovascular and cerebrovascular morbidity and mortality. It is currently unknown whether there are gender-related differences in EPC number and function in middle-aged adults. We tested the hypothesis that EPCs isolated from middle-aged women demonstrate greater colony-forming capacity and migratory activity compared with men of similar age. Peripheral blood samples were collected from 50 sedentary adults, 25 men (59 +/- 1 years of age) and 25 women (58 +/- 1 years of age). Mononuclear cells were isolated and preplated for 2 days, and nonadherent cells were further cultured for 7 days to determine EPC colony-forming units. Migratory activity of EPCs was determined using a modified Boyden chamber. The number of EPC colony-forming units was significantly higher (approximately 150%) in samples collected from women (16 +/- 3) compared with that collected from men (7 +/- 1). In addition, EPC migration (relative fluorescent units) was approximately 40% greater in women (729 +/- 74) than in men (530 +/- 67). In conclusion, these results demonstrate that EPC colony-forming capacity and migratory activity are higher in middle-aged women than in men.

  15. 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

  16. Effect of Lumican on the Migration of Human Mesenchymal Stem Cells and Endothelial Progenitor Cells: Involvement of Matrix Metalloproteinase-14

    PubMed Central

    Perreau, Corinne; Boguslawski, Mateusz; Decot, Véronique; Stoltz, Jean-François; Vallar, Laurent; Niewiarowska, Jolanta; Cierniewski, Czeslaw; Maquart, François-Xavier; Wegrowski, Yanusz; Brézillon, Stéphane

    2012-01-01

    Background Increasing number of evidence shows that soluble factors and extracellular matrix (ECM) components provide an optimal microenvironment controlling human bone marrow mesenchymal stem cell (MSC) functions. Successful in vivo administration of stem cells lies in their ability to migrate through ECM barriers and to differentiate along tissue-specific lineages, including endothelium. Lumican, a protein of the small leucine-rich proteoglycan (SLRP) family, was shown to impede cell migration and angiogenesis. The aim of the present study was to analyze the role of lumican in the control of MSC migration and transition to functional endothelial progenitor cell (EPC). Methodology/Principal Findings Lumican inhibited tube-like structures formation on Matrigel® by MSC, but not EPC. Since matrix metalloproteinases (MMPs), in particular MMP-14, play an important role in remodelling of ECM and enhancing cell migration, their expression and activity were investigated in the cells grown on different ECM substrata. Lumican down-regulated the MMP-14 expression and activity in MSC, but not in EPC. Lumican inhibited MSC, but not EPC migration and invasion. The inhibition of MSC migration and invasion by lumican was reversed by MMP-14 overexpression. Conclusion/Significance Altogether, our results suggest that lumican inhibits MSC tube-like structure formation and migration via mechanisms that involve a decrease of MMP-14 expression and activity. PMID:23236386

  17. Soluble Melanoma Cell Adhesion Molecule (sMCAM/sCD146) Promotes Angiogenic Effects on Endothelial Progenitor Cells through Angiomotin*

    PubMed Central

    Stalin, Jimmy; Harhouri, Karim; Hubert, Lucas; Subrini, Caroline; Lafitte, Daniel; Lissitzky, Jean-Claude; Elganfoud, Nadia; Robert, Stéphane; Foucault-Bertaud, Alexandrine; Kaspi, Elise; Sabatier, Florence; Aurrand-Lions, Michel; Bardin, Nathalie; Holmgren, Lars; Dignat-George, Françoise; Blot-Chabaud, Marcel

    2013-01-01

    The melanoma cell adhesion molecule (CD146) contains a circulating proteolytic variant (sCD146), which is involved in inflammation and angiogenesis. Its circulating level is modulated in different pathologies, but its intracellular transduction pathways are still largely unknown. Using peptide pulldown and mass spectrometry, we identified angiomotin as a sCD146-associated protein in endothelial progenitor cells (EPC). Interaction between angiomotin and sCD146 was confirmed by enzyme-linked immunosorbent assay (ELISA), homogeneous time-resolved fluorescence, and binding of sCD146 on both immobilized recombinant angiomotin and angiomotin-transfected cells. Silencing angiomotin in EPC inhibited sCD146 angiogenic effects, i.e. EPC migration, proliferation, and capacity to form capillary-like structures in Matrigel. In addition, sCD146 effects were inhibited by the angiomotin inhibitor angiostatin and competition with recombinant angiomotin. Finally, binding of sCD146 on angiomotin triggered the activation of several transduction pathways that were identified by antibody array. These results delineate a novel signaling pathway where sCD146 binds to angiomotin to stimulate a proangiogenic response. This result is important to find novel target cells of sCD146 and for the development of therapeutic strategies based on EPC in the treatment of ischemic diseases. PMID:23389031

  18. Contribution of bone marrow-derived endothelial progenitor cells to neovascularization and astrogliosis following spinal cord injury.

    PubMed

    Kamei, Naosuke; Kwon, Sang-Mo; Kawamoto, Atsuhiko; Ii, Masaaki; Ishikawa, Masakazu; Ochi, Mitsuo; Asahara, Takayuki

    2012-12-01

    Spinal cord injury causes initial mechanical damage, followed by ischemia-induced, secondary degeneration, worsening the tissue damage. Although endothelial progenitor cells (EPCs) have been reported to play an important role for pathophysiological neovascularization in various ischemic tissues, the EPC kinetics following spinal cord injury have never been elucidated. In this study, we therefore assessed the in vivo kinetics of bone marrow-derived EPCs by EPC colony-forming assay and bone marrow transplantation from Tie2/lacZ transgenic mice into wild-type mice with spinal cord injury. The number of circulating mononuclear cells and EPC colonies formed by the mononuclear cells peaked at day 3 postspinal cord injury. Bone marrow transplantation study revealed that bone marrow-derived EPCs recruited into the injured spinal cord markedly increased at day 7, when neovascularization and astrogliosis drastically occurred in parallel with axon growth in the damaged tissue. To elucidate further the contribution of EPCs to recovery after spinal cord injury, exogenous EPCs were systemically infused immediately after the injury. The administered EPCs were incorporated into the injured spinal cord and accelerated neovascularization and astrogliosis. These findings suggest that bone marrow-derived EPCs may contribute to the tissue repair by augmenting neovascularization and astrogliosis following spinal cord injury.

  19. 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

  20. Tetrathiomolybdate-associated copper depletion decreases circulating endothelial progenitor cells in women with breast cancer at high risk of relapse

    PubMed Central

    Jain, S.; Cohen, J.; Ward, M. M.; Kornhauser, N.; Chuang, E.; Cigler, T.; Moore, A.; Donovan, D.; Lam, C.; Cobham, M. V.; Schneider, S.; Hurtado Rúa, S. M.; Benkert, S.; Mathijsen Greenwood, C.; Zelkowitz, R.; Warren, J. D.; Lane, M. E.; Mittal, V.; Rafii, S.; Vahdat, L. T.

    2013-01-01

    Background Bone marrow-derived endothelial progenitor cells (EPCs) are critical for metastatic progression. This study explores the effect of tetrathiomolybdate (TM), an anti-angiogenic copper chelator, on EPCs in patients at high risk for breast cancer recurrence. Patients and methods This phase 2 study enrolled breast cancer patients with stage 3 and stage 4 without evidence of disease (NED), and stage 2 if triple-negative. TM 100 mg orally was administered to maintain ceruloplasmin <17 mg/dl for 2 years or until relapse. The primary end point was change in EPCs. Results Forty patients (28 stage 2/3, 12 stage 4 NED) were enrolled. Seventy-five percent patients achieved the copper depletion target by 1 month. Ninety-one percent of triple-negative patients copper-depleted compared with 41% luminal subtypes. In copper-depleted patients only, there was a significant reduction in EPCs/ml by 27 (P = 0.04). Six patients relapsed while on study, of which only one patient had EPCs maintained below baseline. The 10-month relapse-free survival was 85.0% (95% CI 74.6%–96.8%). Only grade 3/4 toxicity was hematologic: neutropenia (3.1% of cycles), febrile neutropenia (0.2%), and anemia (0.2%). Conclusions TM is safe and appears to maintain EPCs below baseline in copper-depleted patients. TM may promote tumor dormancy and ultimately prevent relapse. PMID:23406736

  1. 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.

  2. 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

  3. 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

  4. 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

  5. Different expression of NOS isoforms in early endothelial progenitor cells derived from peripheral and cord blood.

    PubMed

    Muscari, Claudio; Gamberini, Chiara; Carboni, Marco; Basile, Ilaria; Farruggia, Giovanna; Bonafè, Francesca; Giordano, Emanuele; Caldarera, Claudio Marcello; Guarnieri, Carlo

    2007-11-01

    Cord blood and peripheral-adult blood were compared as different sources of early endothelial precursor cells (eEPCs). Total mononuclear cells (MNCs) were obtained from both blood types and committed to eEPCs by exposure to fibronectin, VEGF, IGF-I, and bFGF. Under this condition, MNCs seeded at the density of 3 x 10(5) cells/cm(2) assumed a spindle shape, which was indicative of developing eEPCs, and expanded in a similar manner irrespective to the blood sources. Ulex europaeus agglutinin (UEA-1) and acetylated low density lipoprotein (acLDL) double staining was present in 90% in both peripheral- and cord-blood eEPCs after 2-week expansion. Also, the ability of eEPCs to form tubule-like structures in Matrigel was independent of their blood source, but dependent on the presence of human umbilical vein endothelial cells (HUVECs). eNOS and nNOS were not detectable by Western blotting in both peripheral and cord-blood eEPCs upon 3 weeks and their mRNA levels were lower than 2% relative to those present in HUVECs. On the contrary, iNOS protein was detectable in peripheral-blood eEPCs, but not in cord-blood eEPCs and HUVECs, as well as iNOS mRNA was more concentrated in peripheral-blood eEPCs than in cord-blood eEPCs and HUVECs. These data suggest that: (a) peripheral and cord blood can be considered comparable sources of eEPCs when they are expanded and differentiated in a short-term period; (b) the extremely low expression of constitutive NOS isoforms in the eEPCs of both blood types should markedly reduce their ability to regulate NO-dependent vasorelaxation; (c) the presence of iNOS in peripheral-blood eEPCs could improve the process of vasculogenesis.

  6. Genetic engineering with endothelial nitric oxide synthase improves functional properties of endothelial progenitor cells from patients with coronary artery disease: an in vitro study.

    PubMed

    Kaur, Savneet; Kumar, T R Santhosh; Uruno, Akira; Sugawara, Akira; Jayakumar, Karunakaran; Kartha, Chandrasekharan Cheranellore

    2009-11-01

    Recent studies have reported a marked impairment in the number and functions of endothelial progenitor cells (EPCs) in patients with coronary artery disease (CAD). In view of an important role of eNOS in angiogenesis, in the present study, we evaluated the effects of eNOS gene transfer in ex vivo expanded EPCs isolated from patients with CAD. The expanded EPCs were transfected with mammalian expression vector pcDNA3.1-eNOS containing the full-length human eNOS gene using lipofectamine. About 35-40% of the eNOS-EPCs had higher expression of eNOS as compared to untransfected EPCs. EPCs transfected with pcDNA3.0-EGFP, the plasmid vector expressing green fluorescent protein (GFP) were used as control. The untransfected, GFP-transfected and eNOS-transfected EPCs were compared in terms of important functional attributes of angiogenesis such as proliferation, migration, differentiation and adhesion/integration into tube-like structures in vitro. Functional studies revealed that in the presence of defined growth conditions, compared to the untransfected and GFP-transfected cells, eNOS-EPCs from patients with CAD have a significant increase in [3H] thymidine-labeled DNA (P < 0.01), migration (14.6 +/- 1.8 and 16.5 +/- 1.9 vs. 23.5 +/- 3.4 cells/field, P < 0.01), ability to differentiate into endothelial-like spindle-shaped cells (46 +/- 4.5 and 56.5 +/- 2.1 vs. 93.2 +/- 6.6 cells/field, P < 0.001) and also incorporation into tube-like structures on the matrigel (GFP-EPCs: 21.25 +/- 2.9 vs. GFP-eNOS-EPCs: 34.5 +/- 5.5 cells/field, P < 0.05). We conclude that eNOS gene transfection is a valuable approach to augment angiogenic properties of ex vivo expanded EPCs and eNOS-modified EPCs may offer significant advantages than EPCs alone in terms of their clinical use in patients with myocardial ischemia.

  7. Prognostic Significance of Circulating and Endothelial Progenitor Cell Markers in Type 2 Diabetic Foot

    PubMed Central

    Sambataro, Maria; Seganfreddo, Elena; Canal, Fabio; Furlan, Anna; del Pup, Laura; Niero, Monia; Paccagnella, Agostino; Gherlinzoni, Filippo; dei Tos, Angelo Paolo

    2014-01-01

    Objective. We studied circulating precursor cells (CPC) in type 2 diabetes mellitus (T2DM) with neuropathic foot lesions with or without critical limb ischemia and relationships between endothelial precursor cells (EPC) and peripheral neuropathy. Methods and Subjects. We measured peripheral blood CD34, CD133, and CD45 markers for CPC and KDR, CD31 markers for EPC by citofluorimetry and systemic neural nociceptor CGRP (calcitonin gene related protein) by ELISA in 8 healthy controls (C) and 62 T2DM patients: 14 with neuropathy (N), 20 with neuropathic foot lesions (N1), and 28 with neuroischemic recent revascularized (N2) foot lesions. Timing of lesions was: acute (until 6 weeks), healed, and not healed. Results. CD34+ and CD133+ were reduced in N, N1, and N2 versus C, and CD34+ were lower in N2 versus N1 (P = 0.03). In N2 CD34+KDR+ remain elevated in healed versus chronic lesions and, in N1 CD133+31+ were elevated in acute lesions. CGRP was reduced in N2 and N1 versus C (P < 0.04 versus C 26 ± 2 pg/mL). CD34+KDR+ correlated in N2 with oximetry and negatively in N1 with CGRP. Conclusions. CD34+ CPC are reduced in diabetes with advanced complications and diabetic foot. CD34+KDR+ and CD31+133+ EPC differentiation could have a prognostic and therapeutic significance in the healing process of neuropathic and neuroischemic lesions. PMID:24624298

  8. 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.

  9. 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

  10. 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

  11. MicroRNA-195 regulates proliferation, migration, angiogenesis and autophagy of endothelial progenitor cells by targeting GABARAPL1

    PubMed Central

    Mo, Jianwen; Zhang, Daifen; Yang, Renze

    2016-01-01

    Deep vein thrombosis (DVT) is a common type of venous thrombosis. Successful resolution of DVT-related thrombi is important in the treatment of DVT. Endothelial progenitor cells (EPCs) have emerged as a promising therapeutic choice for DVT-related thrombus resolution; however, the clinical application of EPCs faces many challenges. In the present study, the expression of miR-582, miR-195 and miR-532 under hypoxic or normoxic conditions was measured using quantitative real-time PCR analysis (qRT-PCR) and the results showed that the increased fold of miR-195 was highest in human EPCs (hEPCs) under hypoxic conditions. Then the role and regulating mechanism of miR-195 in improving the function of EPCs was investigated. To investigate the effect of miR-195 inhibition on the autophagy of hEPCs, the expression of the autophagy-related genes LC3B and beclin1 was examined using western blotting, and the formation of autophagosomes was observed using TEM. The results indicated that the inhibition of miR-195 expression could promote autophagy of hEPCs. In addition, we investigated the role of miR-195 on the proliferation, migration and angiogenesis of hEPCs under hypoxia. The results revealed that miR-195 inhibition promotes cell proliferation, migration and angiogenesis of hEPCs under hypoxia. Furthermore, GABA type A receptor associated protein like 1 (GABARAPL1) was identified as a directed target of miR-195 and GABARAPL1 silencing could decrease the effect of miR-195 knockdown on cell proliferation, migration, angiogenesis and autophagy of hEPCs under hypoxia. Together, these results indicate that miR-195 regulates cell proliferation, migration, angiogenesis and autophagy of hEPCs by targeting GABARAPL1. PMID:27623937

  12. 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

  13. Tissue Engineered, Hydrogel-Based Endothelial Progenitor Cell Therapy Robustly Revascularizes Ischemic Myocardium and Preserves Ventricular Function

    PubMed Central

    Atluri, Pavan; Miller, Jordan S; Emery, Robert J; Hung, George; Trubelja, Alen; Cohen, Jeffrey E; Lloyd, Kelsey; Han, Jason; Gaffey, Ann C; MacArthur, John W; Chen, Christopher S; Woo, Y Joseph

    2014-01-01

    Objective Cell based angiogenic therapy for ischemic heart failure has had limited clinical impact, likely related to very low cell retention (<1%) and dispersion. We developed a novel, tissue engineered, hydrogel based cell delivery strategy to overcome these limitations and provide prolonged regional retention of myocardial endothelial progenitor cells (EPC) at high cell dosage. Methods EPCs were isolated from Wistar Rats and encapsulated in fibrin gels. In vitro viability was quantified using a fluorescent live-dead stain of transgenic eGFP+ EPCs. EPC-laden constructs were implanted onto ischemic rat myocardium in a model of acute myocardial infarction (LAD ligation) for 4 weeks. Intramyocardial cell injection (IC, 2×106 EPCs), empty fibrin, and isolated LAD ligation groups served as controls. Hemodynamics were quantified using echocardiography, Doppler flow analysis, and intraventricular pressure-volume analysis. Vasculogenesis and ventricular geometry were quantified. EPC migration was analyzed by utilizing EPCs from transgenic eGFP+ rodents. Results EPCs demonstrated an overall 88.7% viability for all matrix and cell conditions investigated after 48 hours. Histologic assessment of 1-wk implants demonstrated significant migration of transgenic eGFP+ EPCs from the fibrin matrix to the infarcted myocardium as compared to IC (28±12.3 vs. 2.4±2.1cells/hpf, p=0.0001). We also observed a marked increase in vasculogenesis at the implant site. Significant improvements in ventricular hemodynamics and geometry were present following EPC-hydrogel therapy as compared to control. Conclusion We present a tissue engineered hydrogel-based EPC mediated therapy to enhance cell delivery, cell retention, vasculogenesis, and preservation of myocardial structure and function. PMID:25129603

  14. 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

  15. Coenzyme Q10 Attenuates High Glucose-Induced Endothelial Progenitor Cell Dysfunction through AMP-Activated Protein Kinase Pathways.

    PubMed

    Tsai, Hsiao-Ya; Lin, Chih-Pei; Huang, Po-Hsun; Li, Szu-Yuan; Chen, Jia-Shiong; Lin, Feng-Yen; Chen, Jaw-Wen; Lin, Shing-Jong

    2016-01-01

    Coenzyme Q10 (CoQ10), an antiapoptosis enzyme, is stored in the mitochondria of cells. We investigated whether CoQ10 can attenuate high glucose-induced endothelial progenitor cell (EPC) apoptosis and clarified its mechanism. EPCs were incubated with normal glucose (5 mM) or high glucose (25 mM) environment for 3 days, followed by treatment with CoQ10 (10 μM) for 24 hr. Cell proliferation, nitric oxide (NO) production, and JC-1 assay were examined. The specific signal pathways of AMP-activated protein kinase (AMPK), eNOS/Akt, and heme oxygenase-1 (HO-1) were also assessed. High glucose reduced EPC functional activities, including proliferation and migration. Additionally, Akt/eNOS activity and NO production were downregulated in high glucose-stimulated EPCs. Administration of CoQ10 ameliorated high glucose-induced EPC apoptosis, including downregulation of caspase 3, upregulation of Bcl-2, and increase in mitochondrial membrane potential. Furthermore, treatment with CoQ10 reduced reactive oxygen species, enhanced eNOS/Akt activity, and increased HO-1 expression in high glucose-treated EPCs. These effects were negated by administration of AMPK inhibitor. Transplantation of CoQ10-treated EPCs under high glucose conditions into ischemic hindlimbs improved blood flow recovery. CoQ10 reduced high glucose-induced EPC apoptosis and dysfunction through upregulation of eNOS, HO-1 through the AMPK pathway. Our findings provide a potential treatment strategy targeting dysfunctional EPC in diabetic patients.

  16. Senescence Mediated by p16INK4a Impedes Reprogramming of Human Corneal Endothelial Cells into Neural Crest Progenitors

    PubMed Central

    Lu, Wen-Juan; Tseng, Scheffer C. G.; Chen, Shuangling; Tighe, Sean; Zhang, Yuan; Liu, Xin; Chen, Szu-Yu; Su, Chen-Wei; Zhu, Ying-Ting

    2016-01-01

    Human corneal endothelial cells (HCECs) have limited proliferative capacity due to “contact-inhibition” at G1 phase. Such contact-inhibition can be delayed from Day 21 to Day 42 by switching EGF-containing SHEM to LIF/bFGF-containing MESCM through transient activation of LIF-JAK1-STAT3 signaling that delays eventual nuclear translocation of p16INK4a. Using the latter system, we have reported a novel tissue engineering technique by implementing 5 weekly knockdowns with p120 catenin (p120) and Kaiso siRNAs since Day 7 to achieve effective expansion of HCEC monolayers to a transplantable size with a normal HCEC density, through reprogramming of HCECs into neural crest progenitors by activating p120-Kaiso-RhoA-ROCK-canonical BMP signaling. Herein, we noted that a single knockdown with p120-Kaiso siRNAs at Day 42 failed to achieve such reprogramming when contact inhibition transitioned to senescence with nuclear translocation of p16INK4a. In contrast, 5 weekly knockdowns with p120-Kaiso siRNAs since Day 7 precluded senescence mediated by p16INK4a by inducing nuclear translocation of Bmi1 because of sustained activation of JAK2-STAT3 signaling downstream of p120-Kaiso-RhoA-ROCK signaling. STAT3 or Bmi1 siRNA impeded nuclear exclusion of p16INK4a and suppressed the reprogramming induced by p120-Kaiso siRNAs, suggesting that another important engineering strategy of HCEC lies in prevention of senescence mediated by nuclear translocation of p16INK4a. PMID:27739458

  17. C-reactive protein impairs angiogenic functions and decreases the secretion of arteriogenic chemo-cytokines in human endothelial progenitor cells.

    PubMed

    Suh, Wonhee; Kim, Koung Li; Choi, Jin-Ho; Lee, Young-Sam; Lee, Jae-Young; Kim, Jeong-Min; Jang, Hyung-Suk; Shin, In-Soon; Lee, Jung-Sun; Byun, Jonghoe; Jeon, Eun-Seok; Kim, Duk-Kyung

    2004-08-13

    C-reactive protein (CRP), a predictor of future cardiovascular diseases, has been reported to damage the vascular wall by inducing endothelial dysfunction and inflammation. This proatherogenic CRP was speculated to have a role in attenuating angiogenic functions of human endothelial progenitor cells (EPCs), possibly impairing vascular regeneration and increasing cardiovascular vulnerability to ischemic injury. Herein, we investigated the direct effect of CRP on angiogenic activity and gene expression in human EPCs. Incubation of EPCs with human recombinant CRP significantly inhibited EPC migration in response to vascular endothelial growth factor, possibly by decreasing the expression of endothelial nitric oxide synthase and subsequent nitric oxide production. In addition, CRP-treated EPCs showed the reduced adhesiveness onto an endothelial cell monolayer. When assayed for the gene expression of arteriogenic chemo-cytokines, CRP substantially decreased their expression levels in EPC, in part due to the upregulation of suppressors of cytokine signaling proteins. These results suggest that CRP directly attenuates the angiogenic and possibly arteriogenic functions of EPCs. This CRP-induced EPC dysfunction may impair the vascular regenerative capacity of EPCs, thereby leading to increased risk of cardiovascular diseases.

  18. 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.

  19. 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.

  20. 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

  1. 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

  2. Maintaining Moderate Platelet Aggregation and Improving Metabolism of Endothelial Progenitor Cells Increase the Patency Rate of Tissue-Engineered Blood Vessels.

    PubMed

    Wu, Yangxiao; Li, Li; Chen, Wen; Zeng, Wen; Zeng, Lingqin; Wen, Can; Zhu, Chuhong

    2015-07-01

    Small-diameter tissue-engineered blood vessels (TEBVs) have been associated with low, long-term patency rates primarily because of acute thrombosis in early stages and an inability to achieve early endothelialization. Platelets and endothelial progenitor cells (EPCs) play a key role in these processes. A nano delayed-release 5-aminoimidazole-4-carboxamide 1-β-D-ribofuranoside (AICAR)-bound TEBV was implanted in rat carotid arteries for 3 months. AICAR-bound TEBVs had a high patency rate compared with control TEBVs after 3 months. We found that AICAR maintained moderate platelet aggregation in vivo. In vitro data indicated that AICAR inhibits the release of 5-hydroxytryptamine and thromboxane A2 in activating platelets to reduce platelet aggregation. Then, we confirmed that AICAR strengthens the EPC energy state, which results in earlier endothelialization. The homing, migration, and paracrine function of EPCs were enhanced by AICAR in vitro. Besides, AICAR can contribute to the migration of endothelial cells near the anastomosis. The cellularization of TEBVs at different time points was observed too. In conclusion, our study suggests that the application of nanodelivery material containing AICAR can effectively improve small-diameter TEBVs by maintaining moderate platelet aggregation and improving metabolism of EPCs.

  3. 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

  4. Niacin improves ischemia-induced neovascularization in diabetic mice by enhancement of endothelial progenitor cell functions independent of changes in plasma lipids.

    PubMed

    Huang, Po-Hsun; Lin, Chih-Pei; Wang, Chao-Hung; Chiang, Chia-Hung; Tsai, Hsiao-Ya; Chen, Jia-Shiong; Lin, Feng-Yen; Leu, Hsin-Bang; Wu, Tao-Cheng; Chen, Jaw-Wen; Lin, Shing-Jong

    2012-09-01

    Niacin was shown to inhibit acute vascular inflammation and improves endothelial dysfunction independent of changes in plasma lipids. Here, we investigated whether niacin can increase blood flow recovery after tissue ischemia by enhancing endothelial progenitor cell (EPC) functions in diabetic mice. Starting at 4 weeks after the onset of diabetes, vehicle or niacin (40 mg/kg/day) was administered daily by gavage to streptozotocin (STZ)-induced diabetic mice and diabetic endothelial nitric oxide synthase (eNOS)-deficient mice. Unilateral hindlimb ischemia surgery was conducted after 2 weeks of vehicle or niacin treatment. Compared to the control group, the niacin group had significantly increased ischemic/non-ischemic limb blood perfusion ratio and higher capillary density. These effects were markedly reduced in STZ-induced diabetic eNOS-deficient mice. Flow cytometry analysis showed impaired EPC-like cell (Sca-1(+)/Flk-1(+)) mobilization after ischemia surgery in diabetic mice but augmented mobilization in the mice treated with niacin. Diabetes was induced by administering STZ to FVB mice that received eGFP mouse bone marrow cells to evaluate effects of niacin on bone marrow-derived EPC homing and differentiation to endothelial cells. Differentiation of bone marrow-derived EPCs to endothelial cells in the ischemic tissue around vessels in diabetic mice that received niacin treatment, was significantly increased than that in control group. By in vitro studies, incubation with niacin in high-glucose medium reduced H(2)O(2) production, cell apoptosis, and improved high glucose-suppressed EPC functions by nitric oxide-related mechanisms. Our findings demonstrate that niacin increases blood flow recovery after tissue ischemia in diabetic mice through enhancing EPC mobilization and functions via nitric oxide-related pathways.

  5. 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

  6. Granulocyte colony-stimulating factor promotes tumor angiogenesis via increasing circulating endothelial progenitor cells and Gr1+CD11b+ cells in cancer animal models.

    PubMed

    Okazaki, Tatsuma; Ebihara, Satoru; Asada, Masanori; Kanda, Akio; Sasaki, Hidetada; Yamaya, Mutsuo

    2006-01-01

    Recombinant granulocyte colony-stimulating factor (G-CSF) is used for cancer patients with myelosuppression induced by chemotherapy. G-CSF has been reported to progress tumor growth and angiogenesis, but the precise mechanism of tumor angiogenesis activated by G-CSF has not been fully clarified. N-terminal-mutated recombinant human G-CSF administration increased WBCs and neutrophils in peripheral blood and reduced bone marrow stromal cell-derived factor-1 in mice, indicating its biological relevance. Mice were inoculated with Lewis lung carcinoma cells (LLCs) or KLN205 cells and treated with G-CSF. G-CSF accelerated tumor growth and intratumoral vessel density, while it did not accelerate proliferation of LLCs, KLN205 cells or human umbilical vein endothelial cells in vitro. In the absence of tumors, G-CSF did not increase circulating cells that displayed phenotypic characteristics of endothelial progenitor cells (EPCs). In the presence of tumors, G-CSF increased circulating EPCs. In addition, G-CSF treatment increased immune suppressor and endothelial cell-differentiating Gr1+CD11b+ cells in tumor-bearing mice. We conclude that G-CSF promotes tumor growth by activating tumor angiogenesis via increasing circulating EPCs and Gr1+CD11b+ cells in cancer animal models.

  7. 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

  8. 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. PMID:26732848

  9. 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

  10. 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.

  11. 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

  12. 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.

  13. Angiogenic Role of MMP-2/9 Expressed on the Cell Surface of Early Endothelial Progenitor Cells/Myeloid Angiogenic Cells.

    PubMed

    Kanayasu-Toyoda, Toshie; Tanaka, Takeshi; Ishii-Watabe, Akiko; Kitagawa, Hiroko; Matsuyama, Akifumi; Uchida, Eriko; Yamaguchi, Teruhide

    2015-11-01

    Since the introduction of angiogenic cell therapy using early endothelial progenitor cells (EPCs), myeloid angiogenic cells (MACs) have been expected to be useful in treating ischemic diseases. In order to elucidate the angiogenic properties of MACs/EPCs, we clarified the characteristics of MACs as compared to M2 macrophages (Mϕs). Comparison of the gene expression profiles of MACs and late EPCs revealed that MACs expressed greater amounts of metalloproteinase (MMP)-9. It should be noted that the profile of MMP-2/9 expression on the cell surface of MACs was similar to that of M2 Mϕs, and that cell surface MMP-2/9 might be an active form based on molecular size. In addition, the invasion of MACs was prohibited not only by MMP-2/9 inhibitor, but also by the hyaluronidase treatment that caused the down-regulation of MMP-9 on the cell surface of MACs and inhibited their invasion activity. These results indicate that cell surface MMP-2/9 plays an important role in the high invasion ability of MACs. The conditioned medium of both MACs and M2 Mϕs stimulated tube formation of endothelial cells in vitro. MACs caused an increase in vessel formation in in vivo models through the production of IL-8. We propose that the role of MACs with cell surfaces expressing MMP-2/9 is rapidly invading ischemic tissue. PMID:25820539

  14. Vasculoprotective Effects of Combined Endothelial Progenitor Cells and Mesenchymal Stem Cells in Diabetic Wound Care: Their Potential Role in Decreasing Wound-Oxidative Stress

    PubMed Central

    Sukpat, Supakanda; Isarasena, Nipan; Wongphoom, Jutamas; Patumraj, Suthiluk

    2013-01-01

    To investigate whether the combined endothelial progenitor cells (EPCs) and mesenchymal stem cells (MSCs) could enhance angiogenesis and wound healing in diabetic mice. Balb/c nude mice were divided into five groups, including a control group, diabetic group (DM), DM injected with 1 × 106  cells MSCs, DM injected with 1 × 106  cells EPCs, and DM injected with combined 0.5 × 106  cells MSCs and 0.5 × 106  cells EPCs. After seven weeks, the mice were anesthetized, and bilateral full-thickness excision skin wounds were made on the dorsorostral back. The percentage of wound closure in DM group decreased significantly than in control and all other treated groups on day 7 and day 14 (P < 0.005). On day 14, the percentage of capillary vascularity in combine-treated group was significantly higher than in DM (P < 0.005). In the present study, we have demonstrated that the combined EPCs and MSCs can increase vascular endothelial growth factor (VEGF) level and angiogenesis which resulted in reduced neutrophil infiltration, decreased malondialdehyde (MDA) levels, and enhanced wound healing in diabetic mice model. PMID:23844362

  15. Overexpression of osteopontin induces angiogenesis of endothelial progenitor cells via the avβ3/PI3K/AKT/eNOS/NO signaling pathway in glioma cells.

    PubMed

    Wang, Yingyi; Yan, Wei; Lu, Xiaoming; Qian, Chunfa; Zhang, Junxia; Li, Ping; Shi, Lei; Zhao, Peng; Fu, Zhen; Pu, Peiyu; Kang, Chunshen; Jiang, Tao; Liu, Ning; You, Yongping

    2011-08-01

    Angiogenesis, a hallmark of tumor growth, is regulated by various angiogenic factors. Recent studies have shown that osteopontin (OPN) is a secreted, integrin-binding protein that contributes to glioma progression. However, its effect on the angiogenesis of gliomas is not fully understood. To elucidate the role of OPN in the process of glioma angiogenesis, endothelial progenitor cells (EPCs) were treated with conditioned media of human glioma SHG44 cells overexpressing OPN. Here, we identified that OPN secreted by glioma cells accelerated EPCs angiogenesis in vitro, including proliferation, migration, and tube formation. OPN also induced the activation of AKT and endothelial nitric oxide synthase (eNOS) and increased NO production without affecting the expression of VEGF, VEGFR-1, or VEGFR-2. Moreover, the avβ3 antibody, the PI3-K inhibitor LY294002 and the eNOS inhibitor NMA suppressed the OPN-mediated increase in NO production and angiogenesis in EPCs. Taken together, these results demonstrate that OPN directly stimulates angiogenesis via the avβ3/PI3-K/AKT/eNOS/NO signaling pathway and may play an important role in tumorigenesis by enhancing angiogenesis in gliomas.

  16. 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

  17. The Cytoprotective Effects of Human Endothelial Progenitor Cell-Conditioned Medium Against an Ischemic Insult Are Not Dependent on VEGF and IL-8.

    PubMed

    Di Santo, Stefano; Fuchs, Anna-Lena; Periasamy, Ramesh; Seiler, Stefanie; Widmer, Hans Rudolf

    2016-01-01

    Endothelial progenitor cells (EPCs) promote revascularization and tissue repair mainly by paracrine actions. In the present study, we investigated whether EPC-secreted factors in the form of conditioned medium (EPC-CM) can protect cultured brain microvascular endothelial cells against an ischemic insult. Furthermore, we addressed the type of factors that are involved in the EPC-CM-mediated functions. For that purpose, rat brain-derived endothelial cells (rBCEC4 cell line) were exposed to EPC-CM pretreated with proteolytic digestion, heat inactivation, and lipid extraction. Moreover, the involvement of VEGF and IL-8, as canonical angiogenic factors, was investigated by means of neutralizing antibodies. We demonstrated that EPC-CM significantly protected the rBCEC4 cells against an ischemic insult mimicked by induced oxygen-glucose deprivation followed by reoxygenation. The cytoprotective effect was displayed by higher viable cell numbers and reduced caspase 3/7 activity. Heat inactivation, proteolytic digestion, and lipid extraction resulted in a significantly reduced EPC-CM-dependent increase in rBCEC4 viability, tube formation, and survival following the ischemic challenge. Notably, VEGF and IL-8 neutralization did not affect the actions of EPC-CM on rBCEC4 under both standard and ischemic conditions. In summary, our findings show that paracrine factors released by EPCs activate an angiogenic and cytoprotective response on brain microvascular cells and that the activity of EPC-CM relies on the concerted action of nonproteinaceous and proteinaceous factors but do not directly involve VEGF and IL-8. PMID:26776768

  18. Strategies to Enhance the Efficiency of Endothelial Progenitor Cell Therapy by Ephrin B2 Pretreatment and Coadministration with Smooth Muscle Progenitor Cells on Vascular Function During the Wound-Healing Process in Irradiated or Nonirradiated Condition.

    PubMed

    Foubert, Philippe; Squiban, Claire; Holler, Valérie; Buard, Valérie; Dean, Carole; Levy, Bernard I; Benderitter, Marc; Silvestre, Jean Sébastien; Tobelem, Gérard; Tamarat, Radia

    2015-01-01

    Endothelial progenitor cell (EPC) transplantation has beneficial effects for therapeutic neovascularization. We therefore assessed the effect of a therapeutic strategy based on EPC administration in the healing of radiation-induced damage. To improve cell therapy for clinical use, we used pretreatment with ephrin B2-Fc (Eph-B2-Fc) and/or coadministration with smooth muscle progenitor cells. At day 3, EPCs promoted dermal wound healing in both nonirradiated and irradiated mice by 1.2- and 1.15-fold, respectively, compared with animals injected with phosphate-buffered saline. In addition, EPCs also improved skin-blood perfusion and capillary density in both irradiated and nonirradiated mice compared with PBS-injected animals. We also demonstrated that activation with Eph-B2-Fc increased wound closure by 1.6-fold compared with unstimulated EPCs in nonirradiated mice. Interestingly, the beneficial effect of Eph-B2-Fc was abolished in irradiated animals. In addition, we found that Eph-B2-Fc stimulation did not improve EPC-induced vascular permeability or adhesiveness compared to unstimulated EPCs. We hypothesized that this effect was due to high oxidative stress during irradiation, leading to inhibition of EPCs' beneficial effect on vascular function. In this line, we demonstrated that, in irradiated conditions, N-acetyl-l-cysteine treatment restored the beneficial effect of EPC stimulation with Eph-B2-Fc in the wound healing process. In conclusion, stimulation by Eph-B2-Fc improved the beneficial effect of EPCs in physiological conditions and irradiated conditions only in association with antioxidant treatment. Additionally, cotherapy was beneficial in pathological conditions.

  19. 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.

  20. Endothelial cells in co-culture enhance embryonic stem cell differentiation to pancreatic progenitors and insulin-producing cells through BMP signaling.

    PubMed

    Talavera-Adame, Dodanim; Wu, Gordon; He, Yao; Ng, Tina T; Gupta, Ankur; Kurtovic, Silvia; Hwang, Jae Y; Farkas, Daniel L; Dafoe, Donald C

    2011-09-01

    Endothelial cells (ECs) represent the major component of the embryonic pancreatic niche and play a key role in the differentiation of insulin-producing β cells in vivo. However, it is unknown if ECs promote such differentiation in vitro. We investigated whether interaction of ECs with mouse embryoid bodies (EBs) in culture promotes differentiation of pancreatic progenitors and insulin-producing cells and the mechanisms involved. We developed a co-culture system of mouse EBs and human microvascular ECs (HMECs). An increase in the expression of the pancreatic markers PDX-1, Ngn3, Nkx6.1, proinsulin, GLUT-2, and Ptf1a was observed at the interface between EBs and ECs (EB-EC). No expression of these markers was found at the periphery of EBs cultured without ECs or those co-cultured with mouse embryonic fibroblasts (MEFs). At EB-EC interface, proinsulin and Nkx6.1 positive cells co-expressed phospho-Smad1/5/8 (pSmad1/5/8). Therefore, EBs were treated with HMEC conditioned media (HMEC-CM) suspecting soluble factors involved in bone morphogenetic protein (BMP) pathway activation. Upregulation of PDX-1, Ngn3, Nkx6.1, insulin-1, insulin-2, amylin, SUR1, GKS, and amylase as well as down-regulation of SST were detected in treated EBs. In addition, higher expression of BMP-2/-4 and their receptor (BMPR1A) were also found in these EBs. Recombinant human BMP-2 (rhBMP-2) mimicked the effects of the HMEC-CM on EBs. Noggin (NOG), a BMP antagonist, partially inhibited these effects. These results indicate that the differentiation of EBs to pancreatic progenitors and insulin-producing cells can be enhanced by ECs in vitro and that BMP pathway activation is central to this process.

  1. The role of eNOS in the migration and proliferation of bone-marrow derived endothelial progenitor cells and in vitro angiogenesis.

    PubMed

    Lu, Aizhen; Wang, Libo; Qian, Liling

    2015-04-01

    The role of endothelial nitric oxide synthase (eNOS) in the activities of endothelial progenitor cells (EPCs) including migration, proliferation, and tube formation in vitro was investigated. EPCs were obtained from rat bone mononuclear cells by culturing for 7-10 days in EGM-2MV and identified by their capacity for FITC-UEA-1 binding and acetylated low-density lipoprotein (Dil-ac-LDL) intake using fluorescence microscopy. Migration, proliferation and tube formation activities were assessed in the presence or absence of N(ω)-nitro-L-argininemethylester (L-NAME), an eNOS inhibitor. mRNA and protein expression of CXCR4, CXCR7, VEGFR2, and eNOS were detected by real-time PCR and western blotting in the presence or absence of L-NAME. Nitric oxide production was detected by nitrate reductase in the presence or absence of L-NAME. Typical spindle-shaped cells appeared on the 7(th)-10(th) day and confluence reached about 80%. The percentage of FITC-UEA-1 and Dil-ac-LDL double-stained cells was about 85%. Cell migration, proliferation, and tube formation were significantly weakened after eNOS was inhibited (P < 0.05), and the expressions of CXCR4 and eNOS were significantly reduced (P < 0.05, respectively), but there was little change in CXCR7 and VEGFR2. NO production was dramatically decreased after eNOS was inhibited (P < 0.05). In summary, L-NAME significantly reduced the expression of eNOS and NO production by EPCs and inhibited migration, proliferation and tube formation by these cells, suggesting that eNOS affects EPC activities; CXCR4 may be implicated in the action of eNOS.

  2. 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

  3. 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

  4. 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

  5. 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

  6. 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.

  7. 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.

  8. 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

  9. Bradykinin inhibits oxidative stress-induced senescence of endothelial progenitor cells through the B2R/AKT/RB and B2R/EGFR/RB signal pathways

    PubMed Central

    Fu, Cong; Li, Bing; Sun, Yuning; Ma, Genshan; Yao, Yuyu

    2015-01-01

    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

  10. Diabetes-Induced Oxidative Stress in Endothelial Progenitor Cells May Be Sustained by a Positive Feedback Loop Involving High Mobility Group Box-1.

    PubMed

    Wu, Han; Li, Ran; Wei, Zhong-Hai; Zhang, Xin-Lin; Chen, Jian-Zhou; Dai, Qing; Xie, Jun; Xu, Biao

    2016-01-01

    Oxidative stress is considered to be a critical factor in diabetes-induced endothelial progenitor cell (EPC) dysfunction, although the underlying mechanisms are not fully understood. In this study, we investigated the role of high mobility group box-1 (HMGB-1) in diabetes-induced oxidative stress. HMGB-1 was upregulated in both serum and bone marrow-derived monocytes from diabetic mice compared with control mice. In vitro, advanced glycation end productions (AGEs) induced, expression of HMGB-1 in EPCs and in cell culture supernatants in a dose-dependent manner. However, inhibition of oxidative stress with N-acetylcysteine (NAC) partially inhibited the induction of HMGB-1 induced by AGEs. Furthermore, p66shc expression in EPCs induced by AGEs was abrogated by incubation with glycyrrhizin (Gly), while increased superoxide dismutase (SOD) activity in cell culture supernatants was observed in the Gly treated group. Thus, HMGB-1 may play an important role in diabetes-induced oxidative stress in EPCs via a positive feedback loop involving the AGE/reactive oxygen species/HMGB-1 pathway.

  11. 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

  12. 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

  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

    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

  14. EphA2-mediated mesenchymal-amoeboid transition induced by endothelial progenitor cells enhances metastatic spread due to cancer-associated fibroblasts.

    PubMed

    Giannoni, Elisa; Taddei, Maria Letizia; Parri, Matteo; Bianchini, Francesca; Santosuosso, Michela; Grifantini, Renata; Fibbi, Gabriella; Mazzanti, Benedetta; Calorini, Lido; Chiarugi, Paola

    2013-01-01

    Tumor progression is deeply influenced by epigenetic changes induced by tumor stroma. Cancer-associated fibroblasts (CAFs) have been reported to promote epithelial-mesenchymal transition in cancer cells, thereby enhancing their aggressiveness and stem-like properties. As CAFs are able to recruit endothelial progenitor cells (EPCs) to tumor site, we aim to investigate their interplay for prostate carcinoma progression. Both prostate CAFs and cancer cells actively recruit EPCs, known to affect tumor progression through increased vasculogenesis. EPCs synergize with CAFs to further promote epigenetic plasticity of cancer cells, through a mesenchymal-to-amoeboid transition. Indeed, after fibroblasts have engaged epithelial-mesenchymal transition in cancer cells, a further shift towards amoeboid motility is promoted by EPCs through contact-mediated triggering of the bidirectional ephrinA1/EphA2 signaling. The activation of ephrinA1 reverse pathway enhances EPC-induced neo-vascularization, thus promoting tumor growth, while EphA2 forward signaling elicits mesenchymal-amoeboid transition in cancer cells, favoring their adhesion to endothelium, transendothelial migration, and lung metastatic colonization. We therefore underscore that the metastatic advantage given by tumor microenvironment embraces different motility strategies and propose EphA2-targeted tools as useful adjuvants in anti-metastatic treatments.

  15. Functional endothelial progenitor cells selectively recruit neurovascular protective monocyte-derived F4/80(+) /Ly6c(+) macrophages in a mouse model of retinal degeneration.

    PubMed

    Fukuda, Shinichi; Nagano, Masumi; Yamashita, Toshiharu; Kimura, Kenichi; Tsuboi, Ikki; Salazar, Georgina; Ueno, Shinji; Kondo, Mineo; Kunath, Tilo; Oshika, Tetsuro; Ohneda, Osamu

    2013-10-01

    Retinitis pigmentosa is a group of inherited eye disorders that result in profound vision loss with characteristic retinal neuronal degeneration and vasculature attenuation. In a mouse model of retinitis pigmentosa, endothelial progenitor cells (EPC) from bone marrow rescued the vasculature and photoreceptors. However, the mechanisms and cell types underlying these protective effects were uncertain. We divided EPC, which contribute to angiogenesis, into two subpopulations based on their aldehyde dehydrogenase (ALDH) activity and observed that EPC with low ALDH activity (Alde-Low) had greater neuroprotection and vasoprotection capabilities after injection into the eyes of an rd1 mouse model of retinitis pigmentosa compared with EPC with high ALDH activity (Alde-High). Of note, Alde-Low EPC selectively recruited F4/80(+) /Ly6c(+) monocyte-derived macrophages from bone marrow into retina through CCL2 secretion. In addition, the mRNA levels of CCR2, the neurotrophic factors TGF-β1 and IGF-1, and the anti-inflammatory mediator interleukin-10 were higher in migrated F4/80(+) /Ly6c(+) monocyte-derived macrophages as compared with F4/80(+) /Ly6c(-) resident retinal microglial cells. These results suggest a novel therapeutic approach using EPC to recruit neuroprotective macrophages that delay the progression of neural degenerative disease.

  16. 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

  17. 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.

  18. 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

  19. 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.

  20. 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

  1. 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

  2. Recombinant Human Erythropoietin Improves the Neurofunctional Recovery of Rats Following Traumatic Brain Injury via an Increase in Circulating Endothelial Progenitor Cells

    PubMed Central

    Wang, Liang; Wang, Xiaonan; Su, Hua; Han, Zhenying; Yu, Huijie; Wang, Dong; Jiang, Rongcai

    2015-01-01

    Previous studies show that circulating endothelial progenitor cells (EPCs) promote angiogenesis, which is a process associated with improved recovery in animal models of traumatic brain injury (TBI), and that recombinant human erythropoietin (rhEPO) plays a protective role following stroke. Thus, it was hypothesized that rhEPO would enhance recovery following brain injury in a rat model of TBI via an increase in the mobilization of EPCs and, subsequently, in angiogenesis. Flow cytometry assays using CD34− and CD133-specific antibodies were utilized to identify alterations in EPC levels, CD31 and CD34 antibody-stained brain tissue sections were used to quantify angiogenesis, and the Morris water maze (MWM) test and the modified Neurological Severity Score (mNSS) test were used to evaluate behavioral recovery. Compared with saline treatment, treatment with rhEPO significantly increased the number of circulating EPCs on days 1, 4, 7, and 14 (P<0.05), improved spatial learning ability on days 24 and 25 (P<0.05), and enhanced memory recovery on day 26 (P<0.05). Moreover, rhEPO treatment decreased mNSS assessment scores on days 14, 21, and 25 (P<0.05). There was a strong correlation between levels of circulating EPCs and CD34− and CD31-positive cells within the injured boundary zone (CD34+ r=0.910, P<0.01; CD31+ r=0.894, P<0.01) and the ipsilateral hippocampus (CD34+ r=0.841, P<0.01; CD31+ r=0.835, P<0.01). The present data demonstrate that rhEPO treatment improved functional outcomes in rats following TBI via an increase in the mobilization of EPCs and in subsequent angiogenesis. PMID:25085436

  3. 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

  4. 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.

  5. Adherence to lifestyle modifications after a cardiac rehabilitation program and endothelial progenitor cells. A six-month follow-up study.

    PubMed

    Cesari, F; Marcucci, R; Gori, A M; Burgisser, C; Francini, S; Roberts, A T; Sofi, F; Gensini, G F; Abbate, R; Fattirolli, F

    2014-07-01

    An increase of endothelial progenitor cells (EPCs) among acute myocardial infarction (AMI) patients participating in a cardiac rehabilitation (CR) program has been reported, but no data on the impact of adherence to lifestyle recommendations provided during a CR program on EPCs are available. It was our aim to investigate the effect of adherence to lifestyle recommendations on EPCs, inflammatory and functional parameters after six months of a CR program in AMI patients. In 110 AMI patients (90 male/20 female; mean age 57.9 ± 9.4 years) EPCs, high sensitivity C-reactive protein (hsCRP), N-terminal pro-brain natriuretic peptide (NT-ProBNP) levels, and cardiopulmonary testings were determined at the end of the CR (T1) and at a six-month follow-up (T2). At T2 we administered a questionnaire assessing dietary habits and physical activity. At T2, we observed a decrease of EPCs (p<0.05), of hsCRP (p=0.009) and of NT-ProBNP (p<0.0001). Patient population was divided into three categories by Healthy Lifestyle (HL) score (none/low, moderate and high adherence to lifestyle recommendations). We observed a significant association between adherence to lifestyle recommendations, increase in EPCs and exercise capacity between T1 and T2 (Δ EPCs p for trend <0.05; ΔWatt max p for trend=0.004). In a multivariate logistic regression analyses, being in the highest tertile of HL score affected the likelihood of an increase of EPC levels at T2 [OR (95% confidence interval): 3.36 (1.0-10.72) p=0.04]. In conclusion, adherence to lifestyle recommendations provided during a CR program positively influences EPC levels and exercise capacity.

  6. Transfusion of CXCR4-primed endothelial progenitor cells reduces cerebral ischemic damage and promotes repair in db/db diabetic mice.

    PubMed

    Chen, Ji; Chen, Jianying; Chen, Shuzhen; Zhang, Cheng; Zhang, Liangqing; Xiao, Xiang; Das, Avik; Zhao, Yuhui; Yuan, Bin; Morris, Mariana; Zhao, Bin; Chen, Yanfang

    2012-01-01

    This study investigated the role of stromal cell-derived factor-1α (SDF-1α)/CXC chemokine receptor 4 (CXCR4) axis in brain and endothelial progenitor cells (EPCs), and explored the efficacy of CXCR4 primed EPCs in treating ischemic stroke in diabetes. The db/db diabetic and db/+ mice were used in this study. Levels of plasma SDF-1α and circulating CD34+CXCR4+ cells were measured. Brain SDF-1α and CXCR4 expression were quantified at basal and after middle cerebral artery occlusion (MCAO). In in vitro study, EPCs were transfected with adenovirus carrying null (Ad-null) or CXCR4 (Ad-CXCR4) followed with high glucose (HG) treatment for 4 days. For pathway block experiments, cells were pre-incubated with PI3K inhibitor or nitric oxide synthase (NOS) inhibitor for two hours. The CXCR4 expression, function and apoptosis of EPCs were determined. The p-Akt/Akt and p-eNOS/eNOS expression in EPCs were also measured. In in vivo study, EPCs transfected with Ad-null or Ad-CXCR4 were infused into mice via tail vein. On day 2 and 7, the cerebral blood flow, neurologic deficit score, infarct volume, cerebral microvascular density, angiogenesis and neurogenesis were determined. We found: 1) The levels of plasma SDF-1α and circulating CD34+CXCR4+ cells were decreased in db/db mice; 2) The basal level of SDF-1α and MCAO-induced up-regulation of SDF-1α/CXCR4 axis were reduced in the brain of db/db mice; 3) Ad-CXCR4 transfection increased CXCR4 expression in EPCs and enhanced EPC colonic forming capacity; 4) Ad-CXCR4 transfection prevented EPCs from HG-induced dysfunction (migration and tube formation) and apoptosis via activation of PI3K/Akt/eNOS signal pathway; 4) Ad-CXCR4 transfection enhanced the efficacy of EPC infusion in attenuating infarct volume and promoting angiogenesis and neurogenesis. Our data suggest that Ad-CXCR4 primed EPCs have better therapeutic effects for ischemia stroke in diabetes than unmodified EPCs do. PMID:23185548

  7. 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

  8. Nanopatterned acellular valve conduits drive the commitment of blood-derived multipotent cells

    PubMed Central

    Di Liddo, Rosa; Aguiari, Paola; Barbon, Silvia; Bertalot, Thomas; Mandoli, Amit; Tasso, Alessia; Schrenk, Sandra; Iop, Laura; Gandaglia, Alessandro; Parnigotto, Pier Paolo; Conconi, Maria Teresa; Gerosa, Gino

    2016-01-01

    Considerable progress has been made in recent years toward elucidating the correlation among nanoscale topography, mechanical properties, and biological behavior of cardiac valve substitutes. Porcine TriCol scaffolds are promising valve tissue engineering matrices with demonstrated self-repopulation potentiality. In order to define an in vitro model for investigating the influence of extracellular matrix signaling on the growth pattern of colonizing blood-derived cells, we cultured circulating multipotent cells (CMC) on acellular aortic (AVL) and pulmonary (PVL) valve conduits prepared with TriCol method and under no-flow condition. Isolated by our group from Vietnamese pigs before heart valve prosthetic implantation, porcine CMC revealed high proliferative abilities, three-lineage differentiative potential, and distinct hematopoietic/endothelial and mesenchymal properties. Their interaction with valve extracellular matrix nanostructures boosted differential messenger RNA expression pattern and morphologic features on AVL compared to PVL, while promoting on both matrices the commitment to valvular and endothelial cell-like phenotypes. Based on their origin from peripheral blood, porcine CMC are hypothesized in vivo to exert a pivotal role to homeostatically replenish valve cells and contribute to hetero- or allograft colonization. Furthermore, due to their high responsivity to extracellular matrix nanostructure signaling, porcine CMC could be useful for a preliminary evaluation of heart valve prosthetic functionality. PMID:27789941

  9. Endothelial-regenerating cells: an expanding universe.

    PubMed

    Steinmetz, Martin; Nickenig, Georg; Werner, Nikos

    2010-03-01

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

  10. Effects of calcium channel blockers on glucose tolerance, inflammatory state, and circulating progenitor cells in non-diabetic patients with essential hypertension: a comparative study between Azelnidipine and amlodipine on glucose tolerance and endothelial function - a crossover trial (AGENT)

    PubMed Central

    2011-01-01

    Background Hypertension is associated with impaired glucose tolerance and insulin resistance. Medical treatment that interferes with various steps in the renin-angiotensin system improves glucose tolerance and insulin resistance. However, it remains unclear if long-acting calcium channel blockers (CCBs) such as azelnidipine and amlodipine affect glucose tolerance and insulin resistance in clinical practice. Methods Seventeen non-diabetic patients with essential hypertension who had controlled blood pressure levels using amlodipine (5 mg/day) were enrolled in this study. After randomization, either azelnidipine (16 mg/day) or amlodipine (5 mg/day) was administered in a crossover design for 12-weeks. At baseline and the end of each CCB therapy, samples of blood and urine were collected and 75 g oral glucose tolerance test (OGTT) was performed. In addition, hematopoietic progenitor cells (HPCs) were measured at each point by flow cytometry and endothelial functions were measured by fingertip pulse amplitude tonometry using EndoPAT. Results Although blood pressure levels were identical after each CCB treatment, the heart rate significantly decreased after azelnidipine administration than that after amlodipine administration (P < 0.005). Compared with amlodipine administration, azelnidipine significantly decreased levels of glucose and insulin 120 min after the 75 g OGTT (both P < 0.05). Serum levels of high-sensitivity C-reactive protein (P = 0.067) and interleukin-6 (P = 0.035) were decreased. Although endothelial functions were not different between the two medication groups, the number of circulating HPCs was significantly increased after azelnidipine administration (P = 0.016). Conclusions These results suggest that azelnidipine treatment may have beneficial effects on glucose tolerance, insulin sensitivity, the inflammatory state, and number of circulating progenitor cells in non-diabetic patients with essential hypertension. PMID:21906391

  11. Characterization of a Distinct Population of Circulating Human Non-Adherent Endothelial Forming Cells and Their Recruitment via Intercellular Adhesion Molecule-3

    PubMed Central

    Thompson, Emma J.; Barrett, Jeffrey M.; Tooley, Katie; Sen, Shaundeep; Sun, Wai Yan; Grose, Randall; Nicholson, Ian; Levina, Vitalina; Cooke, Ira; Talbo, Gert; Lopez, Angel F.; Bonder, Claudine S.

    2012-01-01

    Circulating vascular progenitor cells contribute to the pathological vasculogenesis of cancer whilst on the other hand offer much promise in therapeutic revascularization in post-occlusion intervention in cardiovascular disease. However, their characterization has been hampered by the many variables to produce them as well as their described phenotypic and functional heterogeneity. Herein we have isolated, enriched for and then characterized a human umbilical cord blood derived CD133+ population of non-adherent endothelial forming cells (naEFCs) which expressed the hematopoietic progenitor cell markers (CD133, CD34, CD117, CD90 and CD38) together with mature endothelial cell markers (VEGFR2, CD144 and CD31). These cells also expressed low levels of CD45 but did not express the lymphoid markers (CD3, CD4, CD8) or myeloid markers (CD11b and CD14) which distinguishes them from ‘early’ endothelial progenitor cells (EPCs). Functional studies demonstrated that these naEFCs (i) bound Ulex europaeus lectin, (ii) demonstrated acetylated-low density lipoprotein uptake, (iii) increased vascular cell adhesion molecule (VCAM-1) surface expression in response to tumor necrosis factor and (iv) in co-culture with mature endothelial cells increased the number of tubes, tubule branching and loops in a 3-dimensional in vitro matrix. More importantly, naEFCs placed in vivo generated new lumen containing vasculature lined by CD144 expressing human endothelial cells (ECs). Extensive genomic and proteomic analyses of the naEFCs showed that intercellular adhesion molecule (ICAM)-3 is expressed on their cell surface but not on mature endothelial cells. Furthermore, functional analysis demonstrated that ICAM-3 mediated the rolling and adhesive events of the naEFCs under shear stress. We suggest that the distinct population of naEFCs identified and characterized here represents a new valuable therapeutic target to control aberrant vasculogenesis. PMID:23144795

  12. Augmentation of Neovascularizaiton in Hindlimb Ischemia by Combined Transplantation of Human Embryonic Stem Cells-Derived Endothelial and Mural Cells

    PubMed Central

    Itoh, Hiroshi; Yamashita, Jun K.; Yurugi-Kobayashi, Takami; Homma, Koichiro; Chao, Ting-Hsing; Miyashita, Kazutoshi; Park, Kwijun; Oyamada, Naofumi; Sawada, Naoya; Taura, Daisuke; Fukunaga, Yasutomo; Tamura, Naohisa; Nakao, Kazuwa

    2008-01-01

    Background We demonstrated that mouse embryonic stem (ES) cells-derived vascular endothelial growth factor receptor-2 (VEGF-R2) positive cells could differentiate into both endothelial cells (EC) and mural cells (MC), and termed them as vascular progenitor cells (VPC). Recently, we have established a method to expand monkey and human ES cells-derived VPC with the proper differentiation stage in a large quantity. Here we investigated the therapeutic potential of human VPC-derived EC and MC for vascular regeneration. Methods and Results After the expansion of human VPC-derived vascular cells, we transplanted these cells to nude mice with hindlimb ischemia. The blood flow recovery and capillary density in ischemic hindlimbs were significantly improved in human VPC-derived EC-transplanted mice, compared to human peripheral and umbilical cord blood-derived endothelial progenitor cells (pEPC and uEPC) transplanted mice. The combined transplantation of human VPC-derived EC and MC synergistically improved blood flow of ischemic hindlimbs remarkably, compared to the single cell transplantations. Transplanted VPC-derived vascular cells were effectively incorporated into host circulating vessels as EC and MC to maintain long-term vascular integrity. Conclusions Our findings suggest that the combined transplantation of human ES cells-derived EC and MC can be used as a new promising strategy for therapeutic vascular regeneration in patients with tissue ischemia. PMID:18301744

  13. Blood-derived DNA methylation markers of cancer risk.

    PubMed

    Marsit, Carmen; Christensen, Brock

    2013-01-01

    The importance of somatic epigenetic alterations in tissues targeted for carcinogenesis is now well recognized and considered a key molecular step in the development of a tumor. Particularly, alteration of gene-specific and genomic DNA methylation has been extensively characterized in tumors, and has become an attractive biomarker of risk due to its specificity and stability in human samples. It also is clear that tumors do not develop as isolated phenomenon in their target tissue, but instead result from altered processes affecting not only the surrounding cells and tissues, but other organ systems, including the immune system. Thus, alterations to DNA methylation profiles detectable in peripheral blood may be useful not only in understanding the carcinogenic process and response to environmental insults, but can also provide critical insights in a systems biological view of tumorigenesis. Research to date has generally focused on how environmental exposures alter genomic DNA methylation content in peripheral blood. More recent work has begun to translate these findings to clinically useful endpoints, by defining the relationship between DNA methylation alterations and cancer risk. This chapter highlights the existing research linking the environment, blood-derived DNA methylation alterations, and cancer risk, and points out how these epigenetic alterations may be contributing fundamentally to carcinogenesis.

  14. Accelerated onset of senescence of endothelial progenitor cells in patients with type 2 diabetes mellitus: role of dimethylarginine dimethylaminohydrolase 2 and asymmetric dimethylarginine.

    PubMed

    Yuan, Qiong; Hu, Chang-Ping; Gong, Zhi-Cheng; Bai, Yong-Ping; Liu, Si-Yu; Li, Yuan-Jian; Jiang, Jun-Lin

    2015-03-20

    The risk of cardiovascular complications in diabetic patients is mainly associated with endothelial dysfunction. Reduced number of EPCs and impaired function of EPCs in diabetes result in imbalance of endothelial homeostasis and dysfunction of vessels. In patients with diabetes mellitus, plasma levels of asymmetric dimethylarginine (ADMA) were elevated, while the expression and activity of dimethylarginine dimethylaminohydrolase (DDAH) were reduced. In the present study, we investigated the role of the DDAH2/ADMA pathway in the senescence of EPCs in type 2 diabetic patients and cultured EPCs treated with high glucose. The results showed that the percentage of senescent EPCs increased while the expression of DDAH2 decreased concomitantly with an increase in the plasma levels of ADMA in patients with type 2 diabetes mellitus (T2DM). Similar results were seen in cultured EPCs treated with high glucose. Exogenous application of ADMA accelerated the senescence of EPCs in a dose-dependent manner, and overexpression of DDAH2 inhibited high glucose-induced EPCs senescence. In addition, it has also been reported that DDAH/ADMA pathway is regulated by silent information regulator 1 (SIRT1) in endothelial cell. In the present study, we found decreased expression of SIRT1 both in T2DM patients and EPCs pretreated with high glucose. And resveratrol (activating SIRT1) inhibited high glucose-induced EPCs senescence by upregulating the expression of DDAH2 and decreasing the levels of ADMA. Taken together, we concluded that DDAH2/ADMA is involved in the accelerated senescence of EPCs in diabetes, which is associated with the activation of SIRT1.

  15. 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

  16. Extracellular matrix-derived products modulate endothelial and progenitor cell migration and proliferation in vitro and stimulate regenerative healing in vivo.

    PubMed

    Vorotnikova, Ekaterina; McIntosh, Donna; Dewilde, Abiche; Zhang, Jianping; Reing, Janet E; Zhang, Li; Cordero, Kevin; Bedelbaeva, Khamilia; Gourevitch, Dimitri; Heber-Katz, Ellen; Badylak, Stephen F; Braunhut, Susan J

    2010-10-01

    Most adult mammals heal without restorative replacement of lost tissue and instead form scar tissue at an injury site. One exception is the adult MRL/MpJ mouse that can regenerate ear and cardiac tissue after wounding with little evidence of scar tissue formation. Following production of a MRL mouse ear hole, 2mm in diameter, a structure rapidly forms at the injury site that resembles the amphibian blastema at a limb amputation site during limb regeneration. We have isolated MRL blastemal cells (MRL-B) from this structure and adapted them to culture. We demonstrate by RT-PCR that even after continuous culturing of these cells they maintain expression of several progenitor cell markers, including DLK (Pref-1), and Msx-1. We have isolated the underlying extracellular matrix (ECM) produced by these MRL-B cells using a new non-proteolytic method and studied the biological activities of this cell-free ECM. Multiplex microELISA analysis of MRL-B cell-free ECM vs. cells revealed selective enrichment of growth factors such as bFGF, HGF and KGF in the matrix compartment. The cell-free ECM, degraded by mild enzyme treatment, was active in promoting migration and proliferation of progenitor cells in vitro and accelerating wound closure in a mouse full thickness cutaneous wound assay in vivo. In vivo, a single application of MRL-B cell matrix-derived products to full thickness cutaneous wounds in non-regenerative mice, B6, induced re-growth of pigmented hair, dermis and epidermis at the wound site whereas scar tissue replaced these tissues at wound sites in mice treated with vehicle alone. These studies suggest that matrix-derived products can stimulate regenerative healing and avert scar tissue formation in adult mammals. PMID:20797438

  17. 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

  18. Progenitor cells in pulmonary vascular remodeling.

    PubMed

    Yeager, Michael E; Frid, Maria G; Stenmark, Kurt R

    2011-01-01

    Pulmonary hypertension is characterized by cellular and structural changes in the walls of pulmonary arteries. Intimal thickening and fibrosis, medial hypertrophy and fibroproliferative changes in the adventitia are commonly observed, as is the extension of smooth muscle into the previously non-muscularized vessels. A majority of these changes are associated with the enhanced presence of α-SM-actin+ cells and inflammatory cells. Atypical abundances of functionally distinct endothelial cells, particularly in the intima (plexiform lesions), and also in the perivascular regions, are also described. At present, neither the origin(s) of these cells nor the molecular mechanisms responsible for their accumulation, in any of the three compartments of the vessel wall, have been fully elucidated. The possibility that they arise from either resident vascular progenitors or bone marrow-derived progenitor cells is now well established. Resident vascular progenitor cells have been demonstrated to exist within the vessel wall, and in response to certain stimuli, to expand and express myofibroblastic, endothelial or even hematopoietic markers. Bone marrow-derived or circulating progenitor cells have also been shown to be recruited to sites of vascular injury and to assume both endothelial and SM-like phenotypes. Here, we review the data supporting the contributory role of vascular progenitors (including endothelial progenitor cells, smooth muscle progenitor cells, pericytes, and fibrocytes) in vascular remodeling. A more complete understanding of the processes by which progenitor cells modulate pulmonary vascular remodeling will undoubtedly herald a renaissance of therapies extending beyond the control of vascular tonicity and reduction of pulmonary artery pressure. PMID:22034593

  19. 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

  20. Progenitor cells in arteriosclerosis: good or bad guys?

    PubMed

    Campagnolo, Paola; Wong, Mei Mei; Xu, Qingbo

    2011-08-15

    Accumulating evidence indicates that the mobilization and recruitment of circulating or tissue-resident progenitor cells that give rise to endothelial cells (ECs) and smooth muscle cells (SMCs) can participate in atherosclerosis, neointima hyperplasia after arterial injury, and transplant arteriosclerosis. It is believed that endothelial progenitor cells do exist and can repair and rejuvenate the arteries under physiologic conditions; however, they may also contribute to lesion formation by influencing plaque stability in advanced atherosclerotic plaque under specific pathologic conditions. At the same time, smooth muscle progenitors, despite their capacity to expedite lesion formation during restenosis, may serve to promote atherosclerotic plaque stabilization by producing extracellular matrix proteins. This profound evidence provides support to the hypothesis that both endothelial and smooth muscle progenitors may act as a double-edged sword in the pathogenesis of arteriosclerosis. Therefore, the understanding of the regulatory networks that control endothelial and smooth muscle progenitor differentiation is undoubtedly fundamental both for basic research and for improving current therapeutic avenues for atherosclerosis. We update the progress in progenitor cell study related to the development of arteriosclerosis, focusing specifically on the role of progenitor cells in lesion formation and discuss the controversial issues that regard the origins, frequency, and impact of the progenitors in the disease.

  1. 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

  2. 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

  3. Neuraminidase enhances in vitro expansion of human erythroid progenitors.

    PubMed

    Bodivit, Gwellaouen; Chadebech, Philippe; Vigon, Isabelle; Yacia, Azzedine; Roziers, Nicolas Burin des; Pirenne, France; Fichelson, Serge

    2016-06-01

    In spite of recent key improvements, in vitro mass production of erythrocytes from human stem cells is still limited by difficulties in obtaining sufficient numbers of erythroid progenitors. In fact, such progenitors are as scarce in the bone marrow as in peripheral blood. We used a two-step culture model of human cord blood-derived erythroid progenitors in the presence or absence of high-purity neuraminidase, in a serum-free, defined culture medium. Granulocytic and megakaryocytic progenitor cell expansions were also studied. We show that significant enhancement of erythroid cell generation is obtained when CD34(+) human hematopoietic progenitors are cultured in the presence of neuraminidase. Interestingly, in so doing, expanded red cell progenitors remained erythropoietin-dependent for further expansion and survival, and cells thus generated displayed a normal phenotype. Moreover, the activity of neuraminidase on these cells can be reversed by simple cell washing. Finally, growth of cells of the other myeloid lineages (granulocytes and megakaryocytes) is either decreased or unchanged in the presence of neuraminidase. This specific feature of neuraminidase, that of stimulation of human red cell progenitor proliferation, provides a safe technique for producing greater numbers of in vitro-generated red blood cells for both basic research and transfusion use. PMID:27478075

  4. Adipose-derived stem cells (ASCs) as a source of endothelial cells in the reconstruction of endothelialized skin equivalents.

    PubMed

    Auxenfans, C; Lequeux, C; Perrusel, E; Mojallal, A; Kinikoglu, B; Damour, O

    2012-07-01

    Tissue-engineered autologous skin is a potential alternative to autograft for burn coverage, but produces poor clinical responses such as unsatisfactory graft intake due to insufficient vascularization. Endothelialized skin equivalents comprising human umbilical vein endothelial cells (HUVECs) survive significantly longer due to inosculation with the capillaries of the host, but these cells are allogeneic by definition. The aim of this study was to reconstruct an autologous endothelialized skin equivalent by incorporating progenitor or pre-differentiated endothelial cells derived from adipose tissue, easily accessible source for autologous transplantation. Human adipose tissue-derived stem cells were isolated from lipoaspirates and amplified to obtain endothelial progenitor cells, which were subsequently differentiated into endothelial cells. These cells were then seeded along with human fibroblasts into a porous collagen-glycosaminoglycan-chitosan scaffold to obtain an endothelialized dermal equivalent. Then, human keratinocytes give rise to a endothelialized skin equivalent. Immunohistochemistry and transmission electron microscopy results demonstrate the presence of capillary-like tubular structures in skin equivalents comprising pre-differentiated endothelial cells, but not endothelial progenitor cells. The former expressed both EN4 and von Willebrand factor, and Weibel-Palade bodies were detected in their cytoplasm. This study demonstrates that adipose tissue is an excellent source of autologous endothelial cells to reconstruct endothelialized tissue equivalents, and that pre-differentiation of stem cells is necessary to obtain vasculature in such models. PMID:21755603

  5. Endothelial colony-forming cells for preparing prevascular three-dimensional cell-dense tissues using cell-sheet engineering.

    PubMed

    Sasagawa, Tadashi; Shimizu, Tatsuya; Yamato, Masayuki; Okano, Teruo

    2016-09-01

    Vascular-derived endothelial cell (EC) network prefabrication in three-dimensional (3D) tissue constructs before transplantation is useful for inducing functional anastomosis with the host vasculature. However, the clinical application of ECs is limited by cell isolation from the existing vasculature, because of the requirement for invasive biopsies and difficulty in obtaining a sufficient number of cells. Endothelial colony-forming cells (ECFCs), which are a subtype of endothelial progenitor cells in the blood, have a strong proliferative and vasculogenic potential. This study attempted to fabricate prevascular 3D cell-dense tissue constructs using cord blood-derived ECFCs and evaluate the in vivo angiogenic potential of these constructs. Human umbilical vascular endothelial cells (HUVECs) were also used in comparison with ECFCs, which were sandwiched between two human dermal-derived fibroblast (FB) sheets using a fibrin-coated cell-sheet manipulator. The inserted ECFCs in double-layered FB sheets were cultured for 3 days, resulting in the formation of network structures similar to those of HUVECs. Additionally, when ECFCs were sandwiched with three FB sheets, a lumen structure was found in the triple-layered cell-sheet constructs at 3 days after co-culture. These constructs containing ECFCs were transplanted into the subcutaneous tissue of immune-deficient rats. One week after transplantation, ECFC-lined functional microvessels containing rat erythrocytes were observed in the same manner as transplanted HUVEC-positive grafts. These results suggest that ECFCs might become an alternative cell source for fabricating a prevascular structure in 3D cell-dense tissue constructs for clinical application. Copyright © 2013 John Wiley & Sons, Ltd.

  6. 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.

  7. Blood-Derived CD4 T Cells Naturally Resist Pyroptosis During Abortive HIV-1 Infection

    PubMed Central

    Muñoz-Arias, Isa; Doitsh, Gilad; Yang, Zhiyuan; Sowinski, Stefanie; Ruelas, Debbie; Greene, Warner C.

    2015-01-01

    Summary Progression to AIDS is driven by CD4 T-cell depletion, mostly involving pyroptosis elicited by abortive HIV infection of CD4 T cells in lymphoid tissues. Inefficient reverse transcription in these cells leads to cytoplasmic accumulation of viral DNAs that are detected by the DNA sensor IFI16, resulting in inflammasome assembly, caspase-1 activation, and pyroptosis. Unexpectedly, we found that peripheral blood-derived CD4 T cells naturally resist pyroptosis. This resistance is partly due to their deeper resting state, resulting in fewer HIV-1 reverse transcripts and lower IFI16 expression. However, when co-cultured with lymphoid-derived cells, blood-derived CD4 T cells become sensitized to pyroptosis, likely recapitulating interactions occurring within lymphoid tissues. Sensitization correlates with higher levels of activated NF-κB, IFI16 expression, and reverse transcription. Blood-derived lymphocytes re-purified from co-cultures lose sensitivity to pyroptosis. These differences highlight how the lymphoid tissue microenvironment encountered by trafficking CD4 T lymphocytes dynamically shapes their biological response to HIV. PMID:26468749

  8. 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

  9. 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

  10. Biodegradable nanoparticle mediated antigen delivery to human cord blood derived dendritic cells for induction of primary T cell responses.

    PubMed

    Diwan, Manish; Elamanchili, Praveen; Lane, Helena; Gainer, Anita; Samuel, John

    2003-01-01

    Dendritic cells (DCs) in the peripheral tissues act as sentinels of the immune system. They detect and capture pathogens entering the body and present their antigens to T cells to trigger responses directed towards elimination of the pathogen. The induction of peripheral tolerance against self and certain foreign antigens is also believed to be mediated through DCs. The outcome of any immune response is largely controlled by the microenvironment of antigen capture, processing and presentation by DCs. The "context" of antigen delivery to DCs will directly influence the microenvironment of antigen presentation and hence the regulation of immune responses. We report here preliminary investigations describing the formulation of a pharmaceutically acceptable, biodegradable, and strategic nanoparticulate delivery system, and its application for efficient antigen loading of DCs to achieve antigen specific T cell activation. "Pathogen-mimicking" nanoparticles capable of interacting with DCs were fabricated by incorporating monophosphoryl lipid A (MPLA; toll-like receptor (TLR) 4 ligand) or CpG ODN (seq #2006; TLR9 ligand) in biodegradable copolymer, poly(D,L,-lactic-co-glycolic acid) (PLGA). The uptake of PLGA nanoparticles by human umbilical cord blood derived DCs (DCs propagated from CD34 progenitors) was conclusively demonstrated by scanning electron microscopy (SEM), fluorescence activated cell sorting (FACS) and confocal laser scanning microscopy (CLSM). Cell phenotype at day 12 of cultures was determined as immature DC using specific cell surface markers, i.e. CD11c (approximately 90%), MHC-II (approximately 70%), CD86 (approximately 20%), CD83 (approximately 5%), CD80 (approximately 40%), CD40 (approximately 40%), and CCR7 (approximately 5%). Tetanus toxoid (TT), a model antigen, was encapsulated in nanoparticles along with an immunomodulator, i.e. either MPLA or CpG ODN. DCs pulsed with various antigen formulations were co-cultured with autologous naïve T cells at

  11. 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

  12. 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

  13. Preserved function of late outgrowth endothelial cells in medically-treated hypertensive patients under well-controlled conditions

    PubMed Central

    Chen, Zhi; Herrmann, Sandra M. S.; Zhu, Xiangyang; Jordan, Kyra L.; Gloviczki, Monika L.; Lerman, Amir; Textor, Stephen C.; Lerman, Lilach O.

    2014-01-01

    Endothelial-progenitor-cells participate in renal repair, but their number and function may be impaired by exposure to cardiovascular risk factors. The number of circulating endothelial-progenitor-cells is decreased in essential and renovascular hypertensive patients, but the effects of hypertension on endothelial-progenitor-cell function are incompletely understood. We hypothesized that endothelial-progenitor-cell function was preserved under well-controlled conditions in treated hypertensive patients. Patients with atherosclerotic-renal-artery-stenosis (n=22) or essential-hypertension (n=24) were studied during controlled sodium intake and anti-hypertensive regimen. Late-outgrowth-endothelial-progenitor-cells were isolated from the inferior vena cava and renal vein blood of atherosclerotic-renal-artery-stenosis and essential-hypertension patients, and a peripheral vein of matched normotensive controls (n=18). The angiogenic function of endothelial-progenitor-cells was assessed in vitro and multi-detector computer tomography used to measure single-kidney hemodynamics and function in atherosclerotic-renal-artery-stenosis and essential-hypertension patients. Inflammatory biomarkers and endothelial-progenitor-cell homing signals levels and renal release were calculated. Inferior vena cava and renal vein-obtained endothelial-progenitor-cell function were similar in atherosclerotic-renal-artery-stenosis and essential-hypertension patients, and comparable to that in normal controls (tube length 171.86±16.846, 191.09±14.222, 174.925±19.774μm, respectively). Function of renal vein-obtained endothelial-progenitor-cells directly correlated with stenotic-kidney glomerular filtration rate, endothelial-progenitor-cell homing factors and anti-inflammatory mediator levels in atherosclerotic-renal-artery-stenosis patients. Therefore, endothelial-progenitor-cell function was relatively preserved in atherosclerotic-renal-artery-stenosis patients, although it directly correlated

  14. 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.

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

    PubMed

    Gendron, Nicolas; Smadja, David M

    2016-08-01

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

  16. The treatment of neurodegenerative disorders using umbilical cord blood and menstrual blood-derived stem cells.

    PubMed

    Sanberg, Paul R; Eve, David J; Willing, Alison E; Garbuzova-Davis, Svitlana; Tan, Jun; Sanberg, Cyndy D; Allickson, Julie G; Cruz, L Eduardo; Borlongan, Cesar V

    2011-01-01

    Stem cell transplantation is a potentially important means of treatment for a number of disorders. Two different stem cell populations of interest are mononuclear umbilical cord blood cells and menstrual blood-derived stem cells. These cells are relatively easy to obtain, appear to be pluripotent, and are immunologically immature. These cells, particularly umbilical cord blood cells, have been studied as either single or multiple injections in a number of animal models of neurodegenerative disorders with some degree of success, including stroke, Alzheimer's disease, amyotrophic lateral sclerosis, and Sanfilippo syndrome type B. Evidence of anti-inflammatory effects and secretion of specific cytokines and growth factors that promote cell survival, rather than cell replacement, have been detected in both transplanted cells.

  17. 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

  18. Human cord blood-derived mononuclear cell transplantation for viral encephalitis-associated cognitive impairment: a case report

    PubMed Central

    2013-01-01

    Introduction Herpes simplex virus is the most common cause of sporadic viral encephalitis. Cognitive impairments persist in most patients who survive herpes simplex virus-caused encephalitis after undergoing currently available treatments. This is the first report on the development of human cord blood-derived mononuclear cell transplantation as a new treatment intervention to improve the prognosis of sequelae of viral encephalitis. Case presentation An 11-year-old Han Chinese boy developed sequelae of viral encephalitis with cognitive, mental and motor impairments in the 8 months following routine treatments. Since receiving allogeneic cord blood-derived mononuclear cell transplantation combined with comprehensive rehabilitation therapies 7 years ago, the patient’s health has significantly improved and remained stable. Conclusions Human cord blood-derived mononuclear cell transplantation may be a potential therapeutic strategy for treating the neuropsychiatric and neurobehavioral sequelae of viral encephalitis. PMID:23835552

  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. [Pulmonary arterial hypertension, bone marrow, endothelial cell precursors and serotonin].

    PubMed

    Ayme-Dietrich, Estelle; Banas, Sophie M; Monassier, Laurent; Maroteaux, Luc

    2016-01-01

    Serotonin and bone-marrow-derived stem cells participate together in triggering pulmonary hypertension. Our work has shown that the absence of 5-HT2B receptors generates permanent changes in the composition of the blood and bone-marrow in the myeloid lineages, particularly in endothelial cell progenitors. The initial functions of 5-HT2B receptors in pulmonary arterial hypertension (PAH) are restricted to bone-marrow cells. They contribute to the differentiation/proliferation/mobilization of endothelial progenitor cells from the bone-marrow. Those bone-marrow-derived cells have a critical role in the development of pulmonary hypertension and pulmonary vascular remodeling. These data indicate that bone-marrow derived endothelial progenitors play a key role in the pathogenesis of PAH and suggest that interactions involving serotonin and bone morphogenic protein type 2 receptor (BMPR2) could take place at the level of the bone-marrow. PMID:27687599

  1. 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

  2. Tumorigenicity Evaluation of Umbilical Cord Blood-derived Mesenchymal Stem Cells.

    PubMed

    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

    2016-07-01

    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.

  3. 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

  4. 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.

  5. The therapeutic potential of human umbilical cord blood-derived mesenchymal stem cells in Alzheimer's disease.

    PubMed

    Lee, Hyun Ju; Lee, Jong Kil; Lee, Hyun; Shin, Ji-woong; Carter, Janet E; Sakamoto, Toshiro; Jin, Hee Kyung; Bae, Jae-sung

    2010-08-30

    The neuropathological hallmarks of Alzheimer's disease (AD) include the presence of extracellular amyloid-beta peptide (Abeta) in the form of amyloid plaques in the brain parenchyma and neuronal loss. The mechanism associated with neuronal death by amyloid plaques is unclear but oxidative stress and glial activation has been implicated. Human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) are being scrutinized as a potential therapeutic tool to prevent various neurodegenerative diseases including AD. However, the therapeutic impact of hUCB-MSCs in AD has not yet been reported. Here we undertook in vitro work to examine the potential impact of hUCB-MSCs treatment on neuronal loss using a paradigm of cultured hippocampal neurons treated with Abeta. We confirmed that hUCB-MSCs co-culture reduced the hippocampal apoptosis induced by Abeta treatment. Moreover, in an acute AD mouse model to directly test the efficacy of hUCB-MSCs treatment on AD-related cognitive and neuropathological outcomes, we demonstrated that markers of glial activation, oxidative stress and apoptosis levels were decreased in AD mouse brain. Interestingly, hUCB-MSCs treated AD mice demonstrated cognitive rescue with restoration of learning/memory function. These data suggest that hUCB-MSCs warrant further investigation as a potential therapeutic agent in AD.

  6. 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

  7. 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

  8. 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

  9. A unique human blood-derived cell population displays high potential for producing insulin.

    PubMed

    Zhao, Yong; Huang, Zhihua; Lazzarini, Ping; Wang, Yong; Di, Anke; Chen, Meiling

    2007-08-17

    Blood can provide a valuable source for the generation of stem cells. Herein we identified a novel cell population from adult human blood, designated peripheral blood insulin-producing cells (PB-IPC). Phenotypic analysis demonstrated that PB-IPC displayed the embryonic stem (ES) cell-associated transcription factors including Oct-4 and Nanog, along with the hematopoietic markers CD9, CD45, and CD117; but lacked expression of the hematopoietic stem cell marker CD34 as well as lymphocyte and monocyte/macrophage markers. Notably, in vitro and in vivo characterization revealed that PB-IPC demonstrated characteristics of islet beta cell progenitors including the expression of beta cell-specific insulin gene transcription factors and prohormone convertases, production of insulin, formation of insulin granules, and the ability to reduce hyperglycemia and migrate into pancreatic islets after transplantation into the diabetic mice. These findings may open up new avenues for autologous blood stem cell-based therapies for diabetes.

  10. Neural progenitor cells regulate microglia functions and activity.

    PubMed

    Mosher, Kira I; Andres, Robert H; Fukuhara, Takeshi; Bieri, Gregor; Hasegawa-Moriyama, Maiko; He, Yingbo; Guzman, Raphael; Wyss-Coray, Tony

    2012-11-01

    We found mouse neural progenitor cells (NPCs) to have a secretory protein profile distinct from other brain cells and to modulate microglial activation, proliferation and phagocytosis. NPC-derived vascular endothelial growth factor was necessary and sufficient to exert at least some of these effects in mice. Thus, neural precursor cells may not only be shaped by microglia, but also regulate microglia functions and activity.

  11. Concomitant lipopolysaccharide-induced transfer of blood-derived components including immunoglobulins into milk.

    PubMed

    Lehmann, M; Wellnitz, O; Bruckmaier, R M

    2013-02-01

    of blood l-lactate concentration. The concomitant changes of all investigated components suggest that they were blood derived. However, the increase in blood components in the milk is not necessarily supportive of the mammary immune system, and likely a side effect of reduced blood-milk barrier integrity.

  12. 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

  13. 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

  14. The Crab Nebula's progenitor

    NASA Technical Reports Server (NTRS)

    Nomoto, K.; Sugimoto, D.; Sparks, W. M.; Fesen, R. A.; Gull, T. R.; Miyaji, S.

    1982-01-01

    The initial mass of the Crab Nebula's progenitor star is estimated by comparing the observed nebular chemical abundances with detailed evolutionary calculations for 2.4- and 2.6-solar-mass helium cores of stars with masses of 8 to 10 solar masses. The results indicate that the mass of the Crab's progenitor was between the upper limit of about 8 solar masses for carbon deflagration and the lower limit of about 9.5 solar masses set by the dredge-up of the helium layer before the development of the helium-burning convective region. A scenario is outlined for the evolution of the progenitor star. It is suggested that the Crab Nebula was probably the product of an electron-capture supernova.

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

    PubMed

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

    2012-09-01

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

  16. Vascular wall progenitor cells in health and disease.

    PubMed

    Psaltis, Peter J; Simari, Robert D

    2015-04-10

    The vasculature plays an indispensible role in organ development and maintenance of tissue homeostasis, such that disturbances to it impact greatly on developmental and postnatal health. Although cell turnover in healthy blood vessels is low, it increases considerably under pathological conditions. The principle sources for this phenomenon have long been considered to be the recruitment of cells from the peripheral circulation and the re-entry of mature cells in the vessel wall back into cell cycle. However, recent discoveries have also uncovered the presence of a range of multipotent and lineage-restricted progenitor cells in the mural layers of postnatal blood vessels, possessing high proliferative capacity and potential to generate endothelial, smooth muscle, hematopoietic or mesenchymal cell progeny. In particular, the tunica adventitia has emerged as a progenitor-rich compartment with niche-like characteristics that support and regulate vascular wall progenitor cells. Preliminary data indicate the involvement of some of these vascular wall progenitor cells in vascular disease states, adding weight to the notion that the adventitia is integral to vascular wall pathogenesis, and raising potential implications for clinical therapies. This review discusses the current body of evidence for the existence of vascular wall progenitor cell subpopulations from development to adulthood and addresses the gains made and significant challenges that lie ahead in trying to accurately delineate their identities, origins, regulatory pathways, and relevance to normal vascular structure and function, as well as disease.

  17. Neurotherapeutic Effect of Cord Blood Derived CD45+ Hematopoietic Cells in Mice after Traumatic Brain Injury

    PubMed Central

    Arien-Zakay, Hadar; Gincberg, Galit; Nagler, Arnon; Cohen, Gadi; Liraz-Zaltsman, Sigal; Trembovler, Victoria; Alexandrovich, Alexander G.; Matok, Ilan; Galski, Hanan; Elchalal, Uriel; Lelkes, Peter I.; Shohami, Esther

    2014-01-01

    Abstract Treatment of traumatic brain injury (TBI) is still an unmet need. Cell therapy by human umbilical cord blood (HUCB) has shown promising results in animal models of TBI and is under evaluation in clinical trials. HUCB contains different cell populations but to date, only mesenchymal stem cells have been evaluated for therapy of TBI. Here we present the neurotherapeutic effect, as evaluated by neurological score, using a single dose of HUCB-derived mononuclear cells (MNCs) upon intravenous (IV) administration one day post-trauma in a mouse model of closed head injury (CHI). Delayed (eight days post-trauma) intracerebroventricular administration of MNCs showed improved neurobehavioral deficits thereby extending the therapeutic window for treating TBI. Further, we demonstrated for the first time that HUCB-derived pan-hematopoietic CD45 positive (CD45+) cells, isolated by magnetic sorting and characterized by expression of CD45 and CD11b markers (96–99%), improved the neurobehavioral deficits upon IV administration, which persisted for 35 days. The therapeutic effect was in a direct correlation to a reduction in the lesion volume and decreased by pre-treatment of the cells with anti-human-CD45 antibody. At the site of brain injury, 1.5-2 h after transplantation, HUCB-derived cells were identified by near infrared scanning and immunohistochemistry using anti-human-CD45 and anti-human-nuclei antibodies. Nerve growth factor and vascular endothelial growth factor levels were differentially expressed in both ipsilateral and contralateral brain hemispheres, thirty-five days after CHI, measured by enzyme-linked immunosorbent assay. These findings indicate the neurotherapeutic potential of HUCB-derived CD45+ cell population in a mouse model of TBI and propose their use in the clinical setting of human TBI. PMID:24640955

  18. 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.

  19. 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.

  20. Stem/Progenitor cells in vascular regeneration.

    PubMed

    Zhang, Li; Xu, Qingbo

    2014-06-01

    A series of studies has been presented in the search for proof of circulating and resident vascular progenitor cells, which can differentiate into endothelial and smooth muscle cells and pericytes in animal and human studies. In terms of pluripotent stem cells, including embryonic stem cells, iPS, and partial-iPS cells, they display a great potential for vascular lineage differentiation. Development of stem cell therapy for treatment of vascular and ischemic diseases remains a major challenging research field. At the present, there is a clear expansion of research into mechanisms of stem cell differentiation into vascular lineages that are tested in animal models. Although there are several clinical trials ongoing that primarily focus on determining the benefits of stem cell transplantation in ischemic heart or peripheral ischemic tissues, intensive investigation for translational aspects of stem cell therapy would be needed. It is a hope that stem cell therapy for vascular diseases could be developed for clinic application in the future.

  1. 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.

  2. 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

  3. 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.

  4. Cord blood-derived CD34+ hematopoietic cells with low mitochondrial mass are enriched in hematopoietic repopulating stem cell function.

    PubMed

    Romero-Moya, Damia; Bueno, Clara; Montes, Rosa; Navarro-Montero, Oscar; Iborra, Francisco J; López, Luis Carlos; Martin, Miguel; Menendez, Pablo

    2013-07-01

    The homeostasis of the hematopoietic stem/progenitor cell pool relies on a fine-tuned balance between self-renewal, differentiation and proliferation. Recent studies have proposed that mitochondria regulate these processes. Although recent work has contributed to understanding the role of mitochondria during stem cell differentiation, it remains unclear whether the mitochondrial content/function affects human hematopoietic stem versus progenitor function. We found that mitochondrial mass correlates strongly with mitochondrial membrane potential in CD34(+) hematopoietic stem/progenitor cells. We, therefore, sorted cord blood CD34(+) cells on the basis of their mitochondrial mass and analyzed the in vitro homeostasis and clonogenic potential as well as the in vivo repopulating potential of CD34(+) cells with high (CD34(+) Mito(High)) versus low (CD34(+) Mito(Low)) mitochondrial mass. The CD34(+) Mito(Low) fraction contained 6-fold more CD34(+)CD38(-) primitive cells and was enriched in hematopoietic stem cell function, as demonstrated by its significantly greater hematopoietic reconstitution potential in immuno-deficient mice. In contrast, the CD34(+) Mito(High) fraction was more enriched in hematopoietic progenitor function with higher in vitro clonogenic capacity. In vitro differentiation of CD34(+) Mito(Low) cells was significantly delayed as compared to that of CD34(+) Mito(High) cells. The eventual complete differentiation of CD34(+) Mito(Low) cells, which coincided with a robust expansion of the CD34(-) differentiated progeny, was accompanied by mitochondrial adaptation, as shown by significant increases in ATP production and expression of the mitochondrial genes ND1 and COX2. In conclusion, cord blood CD34(+) cells with low levels of mitochondrial mass are enriched in hematopoietic repopulating stem cell function whereas high levels of mitochondrial mass identify hematopoietic progenitors. A mitochondrial response underlies hematopoietic stem/progenitor cell

  5. Cord blood-derived CD34+ hematopoietic cells with low mitochondrial mass are enriched in hematopoietic repopulating stem cell function

    PubMed Central

    Romero-Moya, Damia; Bueno, Clara; Montes, Rosa; Navarro-Montero, Oscar; Iborra, Francisco J.; López, Luis Carlos; Martin, Miguel; Menendez, Pablo

    2013-01-01

    The homeostasis of the hematopoietic stem/progenitor cell pool relies on a fine-tuned balance between self-renewal, differentiation and proliferation. Recent studies have proposed that mitochondria regulate these processes. Although recent work has contributed to understanding the role of mitochondria during stem cell differentiation, it remains unclear whether the mitochondrial content/function affects human hematopoietic stem versus progenitor function. We found that mitochondrial mass correlates strongly with mitochondrial membrane potential in CD34+ hematopoietic stem/progenitor cells. We, therefore, sorted cord blood CD34+ cells on the basis of their mitochondrial mass and analyzed the in vitro homeostasis and clonogenic potential as well as the in vivo repopulating potential of CD34+ cells with high (CD34+ MitoHigh) versus low (CD34+ MitoLow) mitochondrial mass. The CD34+ MitoLow fraction contained 6-fold more CD34+CD38− primitive cells and was enriched in hematopoietic stem cell function, as demonstrated by its significantly greater hematopoietic reconstitution potential in immuno-deficient mice. In contrast, the CD34+ MitoHigh fraction was more enriched in hematopoietic progenitor function with higher in vitro clonogenic capacity. In vitro differentiation of CD34+ MitoLow cells was significantly delayed as compared to that of CD34+ MitoHigh cells. The eventual complete differentiation of CD34+ MitoLow cells, which coincided with a robust expansion of the CD34− differentiated progeny, was accompanied by mitochondrial adaptation, as shown by significant increases in ATP production and expression of the mitochondrial genes ND1 and COX2. In conclusion, cord blood CD34+ cells with low levels of mitochondrial mass are enriched in hematopoietic repopulating stem cell function whereas high levels of mitochondrial mass identify hematopoietic progenitors. A mitochondrial response underlies hematopoietic stem/progenitor cell differentiation and proliferation of

  6. 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

  7. 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

  8. Cord Blood Derived CD4+CD25high T Cells Become Functional Regulatory T Cells upon Antigen Encounter

    PubMed Central

    Mayer, Elisabeth; Bannert, Christina; Gruber, Saskia; Klunker, Sven; Spittler, Andreas; Akdis, Cezmi A.

    2012-01-01

    Background: Upon antigen exposure, cord blood derived T cells respond to ubiquitous environmental antigens by high proliferation. To date it remains unclear whether these “excessive” responses relate to different regulatory properties of the putative T regulatory cell (Treg) compartment or even expansion of the Treg compartment itself. Methods: Cord blood (>37 week of gestation) and peripheral blood (healthy controls) were obtained and different Treg cell subsets were isolated. The suppressive potential of Treg populations after antigen exposure was evaluated via functional inhibition assays ([3H]thymidine incorporation assay and CFSE staining) with or without allergen stimulation. The frequency and markers of CD4+CD25highFoxP3+ T cells were characterized by mRNA analysis and flow cytometry. Results: Cord blood derived CD4+CD25high cells did not show substantial suppressor capacity upon TCR activation, in contrast to CD4+CD25high cells freshly purified from adult blood. This could not be explained by a lower frequency of FoxP3+CD4+CD25highcells or FOXP3 mRNA expression. However, after antigen-specific stimulation in vitro, these cells showed strong proliferation and expansion and gained potent suppressive properties. The efficiency of their suppressive capacity can be enhanced in the presence of endotoxins. If T-cells were sorted according to their CD127 expression, a tiny subset of Treg cells (CD4+CD25+CD127low) is highly suppressive even without prior antigen exposure. Conclusion: Cord blood harbors a very small subset of CD4+CD25high Treg cells that requires antigen-stimulation to show expansion and become functional suppressive Tregs. PMID:22272233

  9. 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

  10. 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

  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. 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

    2016-03-08

    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.

  14. 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

  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. 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

  17. Immunohistochemical localization of endothelial cell markers in solitary fibrous tumor.

    PubMed

    Sawada, Namie; Ishiwata, Toshiyuki; Naito, Zenya; Maeda, Shotaro; Sugisaki, Yuichi; Asano, Goro

    2002-12-01

    Solitary fibrous tumor (SFT) is an uncommon tumor first reported in the pleura, but recently described in other tissues. CD34, which is expressed in hematopoietic stem cells, endothelial progenitor cells and vascular endothelial cells, is observed in most SFT and some investigators believe that its expression is a definitive marker of this tumor. In the present study, the expression of vascular endothelial cell markers, such as vascular endothelial growth factor receptor (VEGFR)-1 (flt-1), VEGFR-2 (flk-1/KDR), Tie-2 and c-Met, was examined in SFT to clarify the relationship between SFT and endothelial cells. By immunohistochemical staining of tumor cells from 26 patients, VEGFR-1 was detected in 24 (92%), VEGFR-2 in five (19%), Tie-2 in 14 (54%), and c-Met, a specific receptor of hepatocyte growth factor (HGF) in 23 patients (88%). Furthermore, VEGFR-3 (flt-4) immunoreactivity was detected in eight of 26 patients (31%). In contrast, VEGF, VEGF-C and HGF, which are ligands for the receptors, were not localized in the SFT cells. These findings indicate that most SFT may closely relate to vascular or lymphatic endothelial cells and the endothelial growth factors may contribute to the growth of SFT in a paracrine manner.

  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. Endothelial nitric oxide: protector of a healthy mind.

    PubMed

    Katusic, Zvonimir S; Austin, Susan A

    2014-04-01

    Endothelial nitric oxide (NO) is generated by constitutively active endothelial nitric oxide synthase (eNOS), an essential enzyme responsible for cardiovascular homeostasis. Historically, endothelial NO was first recognized as a major vasodilator involved in control of vasomotor function and local blood flow. In this review, our attention is focused on the emerging role of endothelial NO in linking cerebrovascular function with cognition. We will discuss the recognized ability of endothelial NO to modulate processing of amyloid precursor protein (APP), influence functional status of microglia, and affect cognitive function. Existing evidence suggests that the loss of NO in cultured human cerebrovascular endothelium causes increased expression of APP and β-site APP-cleaving enzyme 1 (BACE1) thereby resulting in increased secretion of amyloid β peptides (Aβ1-40 and Aβ1-42). Furthermore, increased expression of APP and BACE1 as well as increased production of Aβ peptides was detected in the cerebral microvasculature and brain tissue of eNOS-deficient mice. Since Aβ peptides are considered major cytotoxic molecules responsible for the pathogenesis of Alzheimer's disease, these observations support the concept that a loss of endothelial NO might significantly contribute to the initiation and progression of cognitive decline. In addition, genetic inactivation of eNOS causes activation of microglia and promotes a pro-inflammatory phenotype in the brain. Behavioural analysis revealed that eNOS-deficient mice exhibit impaired cognitive performance thereby indicating that selective loss of endothelial NO has a detrimental effect on the function of neuronal cells. Together with findings from prior studies demonstrating the ability of endothelial NO to affect synaptic plasticity, mitochondrial biogenesis, and function of neuronal progenitor cells, it is becoming apparent that the role of endothelial NO in the control of central nervous system function is very complex. We

  20. 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.

  1. 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

  2. Effects of donors' age and passage number on the biological characteristics of menstrual blood-derived stem cells.

    PubMed

    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 differentially-expressed genes associated with age and passage number were identified for use as biomarkers of cell aging.

  3. Electrophysiological characterisation of human umbilical cord blood-derived mesenchymal stem cells induced by olfactory ensheathing cell-conditioned medium.

    PubMed

    Zeng, Yu; Rong, Mingqiang; Liu, Yunsheng; Liu, Jingfang; Lu, Ming; Tao, Xiaoyu; Li, Zhenyan; Chen, Xin; Yang, Kui; Li, Chuntao; Liu, Zhixiong

    2013-12-01

    Umbilical cord blood-derived marrow stromal cells (UCB-MSCs) with high proliferation capacity and immunomodulatory properties are considered to be a good candidate for cell-based therapies. But until now, little work has been focused on the differentiation of UCB-MSCs. In this work, UCB-MSCs were demonstrated to be negative for CD34 and CD45 expression but positive for CD90 and CD105 expression. The gate values of UCB-MSCs for CD90 and CD105 were 99.3 and 98.6 %, respectively. Two weeks after treatment, the percentage of neuron-like cells differentiated from UCB-MSCs was increased to 84 ± 12 % in the experimental group [treated with olfactory ensheathing cells (OECs)-conditioned medium] and they were neuron-specific enolase positive; few neuron-like cells were found in the control group (without OECs-conditioned medium). Using whole-cell recording, sodium and potassium currents were recorded in UCB-MSCs after differentiation by OECs. Thus, human UCB-MSCs could be differentiated to neural cells by secreted secretion from OECs and exhibited electrophysiological properties similar to mature neurons after 2 weeks post-induction. These results imply that OECs can be used as a new strategy for stem cell differentiation and provide an alternative neurogenesis pathway for generating sufficient numbers of neural cells for cell therapy.

  4. Cartilage repair by human umbilical cord blood-derived mesenchymal stem cells with different hydrogels in a rat model.

    PubMed

    Park, Yong-Beom; Song, Minjung; Lee, Choong-Hee; Kim, Jin-A; Ha, Chul-Won

    2015-11-01

    This study was carried out to assess the feasibility of human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) in articular cartilage repair and to further determine a suitable delivering hydrogel in a rat model. Critical sized full thickness cartilage defects were created. The hUCB-MSCs and three different hydrogel composites (hydrogel A; 4% hyaluronic acid/30% pluronic (1:1, v/v), hydrogel B; 4% hyaluronic acid, and hydrogel C; 4% hyaluronic acid/30% pluronic/chitosan (1:1:2, v/v)) were implanted into the experimental knee (right knee) and hydrogels without hUCB-MSCs were implanted into the control knee (left knee). Defects were evaluated after 8 weeks. The hUCB-MSCs with hydrogels composites resulted in a better repair as seen by gross and histological evaluation compared with hydrogels without hUCB-MSCs. Among the three different hydrogels, the 4% hyaluronic acid hydrogel composite (hydrogel B) showed the best result in cartilage repair as seen by the histological evaluation compared with the other hydrogel composites (hydrogel A and C). The results of this study suggest that hUCB-MSCs may be a promising cell source in combination with 4% hyaluronic acid hydrogels in the in vivo repair of cartilage defects.

  5. 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

  6. Effects of donors' age and passage number on the biological characteristics of menstrual blood-derived stem cells.

    PubMed

    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 differentially-expressed genes associated with age and passage number were identified for use as biomarkers of cell aging. PMID:26823782

  7. 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

  8. [Vascular endothelial Barrier Function].

    PubMed

    Ivanov, A N; Puchinyan, D M; Norkin, I A

    2015-01-01

    Endothelium is an important regulator of selective permeability of the vascular wall for different molecules and cells. This review summarizes current data on endothelial barrier function. Endothelial glycocalyx structure, its function and role in the molecular transport and leukocytes migration across the endothelial barrier are discussed. The mechanisms of transcellular transport of macromolecules and cell migration through endothelial cells are reviewed. Special section of this article addresses the structure and function of tight and adherens endothelial junction, as well as their importance for the regulation of paracellular transport across the endothelial barrier. Particular attention is paid to the signaling mechanism of endothelial barrier function regulation and the factors that influence on the vascular permeability.

  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 concentration of ethanol favors progenitor cell differentiation and neovascularization in high-fat diet-fed mice model.

    PubMed

    Vergori, Luisa; Lauret, Emilie; Soleti, Raffaella; Martinez, Maria Carmen; Andriantsitohaina, Ramaroson

    2016-09-01

    Endothelial progenitor cells (EPCs) and monocytic cells from bone marrow (BM) can be recruited to the injured endothelium and contribute to its regeneration. During metabolic diseases such as obesity and diabetes, progenitor cell function is impaired. Several studies have shown that moderate alcohol consumption prevents the development and progression of atherosclerosis in a variety of animal/mouse models and increases mobilization of progenitor cells. Along with these studies, we identify ethanol at low concentration as therapeutic tool to in vitro expand progenitor cells in order to obtain an adequate number of cells for their use in the treatment of cardiovascular diseases. We evaluated the effects of ethanol on the phenotype of BM-derived cells from mice fed with high-fat diet (HFD). HFD did not induce changes in weight of mice but induced metabolic alterations. HFD feeding increased the differentiation of monocytic progenitors but not EPCs. Whereas ethanol at 0.6% is able to increase monocytic progenitor differentiation, 1% ethanol diminished it. Furthermore, ethanol at 0.6% increased the ability of progenitor cells to promote in vivo angiogenesis as well as secretome of BM-derived cells from mice fed with HFD, but not in mice fed normal diet. In conclusion, ethanol at low concentration is able to increase angiogenic abilities of progenitor cells from animals with early metabolic alterations.

  12. 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

  13. Stem and progenitor cells: advancing bone tissue engineering.

    PubMed

    Tevlin, R; Walmsley, G G; Marecic, O; Hu, Michael S; Wan, D C; Longaker, M T

    2016-04-01

    Unlike many other postnatal tissues, bone can regenerate and repair itself; nevertheless, this capacity can be overcome. Traditionally, surgical reconstructive strategies have implemented autologous, allogeneic, and prosthetic materials. Autologous bone--the best option--is limited in supply and also mandates an additional surgical procedure. In regenerative tissue engineering, there are myriad issues to consider in the creation of a functional, implantable replacement tissue. Importantly, there must exist an easily accessible, abundant cell source with the capacity to express the phenotype of the desired tissue, and a biocompatible scaffold to deliver the cells to the damaged region. A literature review was performed using PubMed; peer-reviewed publications were screened for relevance in order to identify key advances in stem and progenitor cell contribution to the field of bone tissue engineering. In this review, we briefly introduce various adult stem cells implemented in bone tissue engineering such as mesenchymal stem cells (including bone marrow- and adipose-derived stem cells), endothelial progenitor cells, and induced pluripotent stem cells. We then discuss numerous advances associated with their application and subsequently focus on technological advances in the field, before addressing key regenerative strategies currently used in clinical practice. Stem and progenitor cell implementation in bone tissue engineering strategies have the ability to make a major impact on regenerative medicine and reduce patient morbidity. As the field of regenerative medicine endeavors to harness the body's own cells for treatment, scientific innovation has led to great advances in stem cell-based therapies in the past decade.

  14. Mesenchymal progenitor cells for the osteogenic lineage

    PubMed Central

    Ono, Noriaki; Kronenberg, Henry M.

    2015-01-01

    Mesenchymal progenitors of the osteogenic lineage provide the flexibility for bone to grow, maintain its function and homeostasis. Traditionally, colony-forming-unit fibroblasts (CFU-Fs) have been regarded as surrogates for mesenchymal progenitors; however, this definition cannot address the function of these progenitors in their native setting. Transgenic murine models including lineage-tracing technologies based on the cre-lox system have proven to be useful in delineating mesenchymal progenitors in their native environment. Although heterogeneity of cell populations of interest marked by a promoter-based approach complicates overall interpretation, an emerging complexity of mesenchymal progenitors has been revealed. Current literatures suggest two distinct types of bone progenitor cells; growth-associated mesenchymal progenitors contribute to explosive growth of bone in early life, whereas bone marrow mesenchymal progenitors contribute to the much slower remodeling process and response to injury that occurs mainly in adulthood. More detailed relationships of these progenitors need to be studied through further experimentation. PMID:26526380

  15. Good manufacturing practice-compliant isolation and culture of human umbilical cord blood-derived mesenchymal stem cells

    PubMed Central

    2014-01-01

    Background Mesenchymal stem cells (MSCs) are an attractive source of stem cells for clinical applications. These cells exhibit a multilineage differentiation potential and strong capacity for immune modulation. Thus, MSCs are widely used in cell therapy, tissue engineering, and immunotherapy. Because of important advantages, umbilical cord blood-derived MSCs (UCB-MSCs) have attracted interest for some time. However, the applications of UCB-MSCs are limited by the small number of recoverable UCB-MSCs and fetal bovine serum (FBS)-dependent expansion methods. Hence, this study aimed to establish a xenogenic and allogeneic supplement-free expansion protocol. Methods UCB was collected to prepare activated platelet-rich plasma (aPRP) and mononuclear cells (MNCs). aPRP was applied as a supplement in Iscove modified Dulbecco medium (IMDM) together with antibiotics. MNCs were cultured in complete IMDM with four concentrations of aPRP (2, 5, 7, or 10%) or 10% FBS as the control. The efficiency of the protocols was evaluated in terms of the number of adherent cells and their expansion, the percentage of successfully isolated cells in the primary culture, surface marker expression, and in vitro differentiation potential following expansion. Results The results showed that primary cultures with complete medium containing 10% aPRP exhibited the highest success, whereas expansion in complete medium containing 5% aPRP was suitable. UCB-MSCs isolated using this protocol maintained their immunophenotypes, multilineage differentiation potential, and did not form tumors when injected at a high dose into athymic nude mice. Conclusion This technique provides a method to obtain UCB-MSCs compliant with good manufacturing practices for clinical application. PMID:24565047

  16. 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.

  17. [Influence of different gelatin concentration and lymphocyte isolation liquid on primary culture of umbilical cord blood derived adhesive cells].

    PubMed

    Zhang, Cheng; Chen, Xing-Hua; Zhang, Xi; Gao, Lei; Kong, Pei-Yan; Liu, Hong; Liang, Xue; Peng, Xian-Gui; Wang, Qing-Yu

    2008-12-01

    In order to study the influence of different gelatin concentrations, and lymphocyte isolation liquid on primary culture of umbilical cord blood-derived adhesive cells (hCBACs), the red blood cells of umbilical cord blood was separated by 3% and 6 % gelatin for detecting the effectiveness of sedimentation, then the adhesion rate at 48 hours, the day of initial expansion and the rate of culture success were detected for hCBACs cultured with CD34(+) cells after the mononuclear cells were separated by 6% gelatin followed by Ficoll and Percoll, and the morphological characteristics and growth status were observed by invert microscopy. Cytochemistry stain for nonspecific esterase stain (NSE), peroxidase (POX), periodic acid Schiff reaction (PAS) and alkali phosphatase (ALP) and immunocytochemistry labeling for CD31, CD45, CD68 and fibronectin (Fn) were detected. The results showed that 6 % gelatin was better than that 3% gelatin for red blood sedimentation. The Percoll was predominant over Ficoll in adhesion rate at 48 hours, the day of initial expansion, the time of initial formation of adhesive cell colony units, the time of maximal numbers of adhesive cell colony units, the the cell fusion time and ratio of culture success. 60% fibroblast-liked cells, 36% macrophage liked cells and 4% small-round cells were observed in cells isolated by both isolated methods. The cytochemistry stain for NSE, POX, PAS and ALP was similar in two groups, the difference was not statistically significant between these two groups. The immunocytochemistry labeling for CD31, CD45, CD68 and Fn was also similar in both groups and the difference was also not statistically significant between these two groups. It is concluded that the combination of 6% gelatin with Percoll is an ideal separation method for primary culture of hCBACs, which provides basic information for clinical application.

  18. Inducible HGF-secreting Human Umbilical Cord Blood-derived MSCs Produced via TALEN-mediated Genome Editing Promoted Angiogenesis.

    PubMed

    Chang, Hyun-Kyung; Kim, Pyung-Hwan; Cho, Hyun-Min; Yum, Soo-Young; Choi, Young-Jin; Son, YeonSung; Lee, DaBin; Kang, InSung; Kang, Kyung-Sun; Jang, Goo; Cho, Je-Yoel

    2016-09-01

    Mesenchymal stem cells (MSCs) promote therapeutic angiogenesis to cure serious vascular disorders. However, their survival period and cytokine-secretory capacity are limited. Although hepatocyte growth factor (HGF) can accelerate the rate of angiogenesis, recombinant HGF is limited because of its very short half-life (<3-5 minutes). Thus, continuous treatment with HGF is required to obtain an effective therapeutic response. To overcome these limitations, we produced genome-edited MSCs that secreted HGF upon drug-specific induction. The inducible HGF expression cassette was integrated into a safe harbor site in an MSC chromosome using the TALEN system, resulting in the production of TetOn-HGF/human umbilical cord blood-derived (hUCB)-MSCs. Functional assessment of the TetOn-HGF/hUCB-MSCs showed that they had enhanced mobility upon the induction of HGF expression. Moreover, long-term exposure by doxycycline (Dox)-treated TetOn-HGF/hUCB-MSCs enhanced the anti-apoptotic responses of genome-edited MSCs subjected to oxidative stress and improved the tube-formation ability. Furthermore, TetOn-HGF/hUCB-MSCs encapsulated by arginine-glycine-aspartic acid (RGD)-alginate microgel induced to express HGF improved in vivo angiogenesis in a mouse hindlimb ischemia model. This study showed that the inducible HGF-expressing hUCB-MSCs are competent to continuously express and secrete HGF in a controlled manner. Thus, the MSCs that express HGF in an inducible manner are a useful therapeutic modality for the treatment of vascular diseases requiring angiogenesis.

  19. 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

  20. Expression of Coxsackievirus and Adenovirus Receptor Separates Hematopoietic and Cardiac Progenitor Cells in Fetal Liver Kinase 1-Expressing Mesoderm

    PubMed Central

    Tashiro, Katsuhisa; Hirata, Nobue; Okada, Atsumasa; Yamaguchi, Tomoko; Takayama, Kazuo; Mizuguchi, Hiroyuki

    2015-01-01

    In developing embryos or in vitro differentiation cultures using pluripotent stem cells (PSCs), such as embryonic stem cells and induced pluripotent stem cells, fetal liver kinase 1 (Flk1)-expressing mesodermal cells are thought to be a heterogeneous population that includes hematopoietic progenitors, endothelial progenitors, and cardiac progenitors. However, information on cell surface markers for separating these progenitors in Flk1+ cells is currently limited. In the present study, we show that distinct types of progenitor cells in Flk1+ cells could be separated according to the expression of coxsackievirus and adenovirus receptor (CAR, also known as CXADR), a tight junction component molecule. We found that mouse and human PSC- and mouse embryo-derived Flk1+ cells could be subdivided into Flk1+CAR+ cells and Flk1+CAR− cells. The progenitor cells with cardiac potential were almost entirely restricted to Flk1+CAR+ cells, and Flk1+CAR− cells efficiently differentiated into hematopoietic cells. Endothelial differentiation potential was observed in both populations. Furthermore, from the expression of CAR, Flk1, and platelet-derived growth factor receptor-α (PDGFRα), Flk1+ cells could be separated into three populations (Flk1+PDGFRα−CAR− cells, Flk1+PDGFRα−CAR+ cells, and Flk1+PDGFRα+CAR+ cells). Flk1+PDGFRα+ cells and Flk1+PDGFRα− cells have been reported as cardiac and hematopoietic progenitor cells, respectively. We identified a novel population (Flk1+PDGFRα−CAR+ cells) with the potential to differentiate into not only hematopoietic cells and endothelial cells but also cardiomyocytes. Our findings indicate that CAR would be a novel and prominent marker for separating PSC- and embryo-derived Flk1+ mesodermal cells with distinct differentiation potentials. PMID:25762001

  1. 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

  2. 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.

  3. 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

  4. Molecular properties of adult mouse gastric and intestinal epithelial progenitors in their niches.

    PubMed

    Giannakis, Marios; Stappenbeck, Thaddeus S; Mills, Jason C; Leip, Douglas G; Lovett, Michael; Clifton, Sandra W; Ippolito, Joseph E; Glasscock, Jarret I; Arumugam, Manimozhiyan; Brent, Michael R; Gordon, Jeffrey I

    2006-04-21

    We have sequenced 36,641 expressed sequence tags from laser capture microdissected adult mouse gastric and small intestinal epithelial progenitors, obtaining 4031 and 3324 unique transcripts, respectively. Using Gene Ontology (GO) terms, each data set was compared with cDNA libraries from intact adult stomach and small intestine. Genes in GO categories enriched in progenitors were filtered against genes in GO categories represented in hematopoietic, neural, and embryonic stem cell transcriptomes and mapped onto transcription factor networks, plus canonical signal transduction and metabolic pathways. Wnt/beta-catenin, phosphoinositide-3/Akt kinase, insulin-like growth factor-1, vascular endothelial growth factor, integrin, and gamma-aminobutyric acid receptor signaling cascades, plus glycerolipid, fatty acid, and amino acid metabolic pathways are among those prominently represented in adult gut progenitors. The results reveal shared as well as distinctive features of adult gut stem cells when compared with other stem cell populations.

  5. [Endothelial cell adhesion molecules].

    PubMed

    Ivanov, A N; Norkin, I A; Puchin'ian, D M; Shirokov, V Iu; Zhdanova, O Iu

    2014-01-01

    The review presents current data concerning the functional role of endothelial cell adhesion molecules belonging to different structural families: integrins, selectins, cadherins, and the immunoglobulin super-family. In this manuscript the regulatory mechanisms and factors of adhesion molecules expression and distribution on the surface of endothelial cells are discussed. The data presented reveal the importance of adhesion molecules in the regulation of structural and functional state of endothelial cells in normal conditions and in pathology. Particular attention is paid to the importance of these molecules in the processes of physiological and pathological angiogenesis, regulation of permeability of the endothelial barrier and cell transmigration.

  6. 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.

  7. Erythrocyte enrichment in hematopoietic progenitor cell cultures based on magnetic susceptibility of the hemoglobin.

    PubMed

    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.

  8. 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

  9. 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 understanding of ARDS and lead to development of therapeutics. This review 1) summarizes the advantages and pitfalls of EC cultures to study vascular leak in ARDS, 2) provides an overview of elements of the microvascular environment that can directly affect endothelial barrier function, and 3) discusses alternative methods to bridge the gap between basic research and clinical application with the intent of improving the translational value of present EC culture approaches. PMID:27343194

  10. Impact of Charged Particle Exposure on Homologous DNA Double-Strand Break Repair in Human Blood-Derived Cells

    PubMed Central

    Rall, Melanie; Kraft, Daniela; Volcic, Meta; Cucu, Aljona; Nasonova, Elena; Taucher-Scholz, Gisela; Bönig, Halvard; Wiesmüller, Lisa; Fournier, Claudia

    2015-01-01

    Ionizing radiation generates DNA double-strand breaks (DSB) which, unless faithfully repaired, can generate chromosomal rearrangements in hematopoietic stem and/or progenitor cells (HSPC), potentially priming the cells towards a leukemic phenotype. Using an enhanced green fluorescent protein (EGFP)-based reporter system, we recently identified differences in the removal of enzyme-mediated DSB in human HSPC versus mature peripheral blood lymphocytes (PBL), particularly regarding homologous DSB repair (HR). Assessment of chromosomal breaks via premature chromosome condensation or γH2AX foci indicated similar efficiency and kinetics of radiation-induced DSB formation and rejoining in PBL and HSPC. Prolonged persistence of chromosomal breaks was observed for higher LET charged particles which are known to induce more complex DNA damage compared to X-rays. Consistent with HR deficiency in HSPC observed in our previous study, we noticed here pronounced focal accumulation of 53BP1 after X-ray and carbon ion exposure (intermediate LET) in HSPC versus PBL. For higher LET, 53BP1 foci kinetics was similarly delayed in PBL and HSPC suggesting similar failure to repair complex DNA damage. Data obtained with plasmid reporter systems revealed a dose- and LET-dependent HR increase after X-ray, carbon ion and higher LET exposure, particularly in HR-proficient immortalized and primary lymphocytes, confirming preferential use of conservative HR in PBL for intermediate LET damage repair. HR measured adjacent to the leukemia-associated MLL breakpoint cluster sequence in reporter lines revealed dose dependency of potentially leukemogenic rearrangements underscoring the risk of leukemia-induction by radiation treatment. PMID:26618143

  11. Circulating endothelial cells and angiogenic serum factors during neoadjuvant chemotherapy of primary breast cancer

    PubMed Central

    Fürstenberger, G; von Moos, R; Lucas, R; Thürlimann, B; Senn, H-J; Hamacher, J; Boneberg, E-M

    2006-01-01

    Circulating endothelial cells (CECs) as well as bone-marrow-derived endothelial precursor cells (EPC) play an important role in neovascularisation and tumour growth. To study the impact of neoadjuvant chemotherapy on the amounts of CEC and their precursor cells, mature CEC and their progenitors were quantified by flow cytometry in peripheral blood of breast cancer patients during anthracycline and/or taxane based neoadjuvant chemotherapy and subsequent surgery in comparison to age-matched healthy controls. Cell numbers were tested for correlation with serum levels of angiopoietin-2, erythropoietin, endostatin, endoglin, VEGF and sVCAM-1 as well as clinical and pathological features of breast cancer disease. Circulating endothelial cells were significantly elevated in breast cancer patients and decreased during chemotherapy, whereas EPC (CD34+/VEGFR-2+) as well as their progenitor cell population CD133+/CD34+ and the population of CD34+ stem cells increased. Concomitantly with the increase of progenitor cells an increase of VEGF, erythropoietin and angiopoietin-2 was observed. These data suggest that chemotherapy can only reduce the amounts of mature CEC, probably reflecting detached cells from tumour vessels, whereas the EPC and their progenitors are mobilised by chemotherapy. Since this mobilisation of EPC may contribute to tumour neovascularisation an early antiangiogenic therapy in combination with chemotherapy could be beneficial for the success of cancer therapy. PMID:16450002

  12. HDL and endothelial protection

    PubMed Central

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

    2013-01-01

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

  13. PROGENITORS OF RECOMBINING SUPERNOVA REMNANTS

    SciTech Connect

    Moriya, Takashi J.

    2012-05-01

    Usual supernova remnants have either ionizing plasma or plasma in collisional ionization equilibrium, i.e., the ionization temperature is lower than or equal to the electron temperature. However, the existence of recombining supernova remnants, i.e., supernova remnants with ionization temperature higher than the electron temperature, has been recently confirmed. One suggested way to have recombining plasma in a supernova remnant is to have a dense circumstellar medium at the time of the supernova explosion. If the circumstellar medium is dense enough, collisional ionization equilibrium can be established in the early stage of the evolution of the supernova remnant and subsequent adiabatic cooling, which occurs after the shock wave gets out of the dense circumstellar medium, makes the electron temperature lower than the ionization temperature. We study the circumstellar medium around several supernova progenitors and show which supernova progenitors can have a circumstellar medium dense enough to establish collisional ionization equilibrium soon after the explosion. We find that the circumstellar medium around red supergiants (especially massive ones) and the circumstellar medium dense enough to make Type IIn supernovae can establish collisional ionization equilibrium soon after the explosion and can evolve to become recombining supernova remnants. Wolf-Rayet stars and white dwarfs have the possibility to be recombining supernova remnants but the fraction is expected to be very small. As the occurrence rate of the explosions of red supergiants is much higher than that of Type IIn supernovae, the major progenitors of recombining supernova remnants are likely to be red supergiants.

  14. Progenitors of Recombining Supernova Remnants

    NASA Astrophysics Data System (ADS)

    Moriya, Takashi J.

    2012-05-01

    Usual supernova remnants have either ionizing plasma or plasma in collisional ionization equilibrium, i.e., the ionization temperature is lower than or equal to the electron temperature. However, the existence of recombining supernova remnants, i.e., supernova remnants with ionization temperature higher than the electron temperature, has been recently confirmed. One suggested way to have recombining plasma in a supernova remnant is to have a dense circumstellar medium at the time of the supernova explosion. If the circumstellar medium is dense enough, collisional ionization equilibrium can be established in the early stage of the evolution of the supernova remnant and subsequent adiabatic cooling, which occurs after the shock wave gets out of the dense circumstellar medium, makes the electron temperature lower than the ionization temperature. We study the circumstellar medium around several supernova progenitors and show which supernova progenitors can have a circumstellar medium dense enough to establish collisional ionization equilibrium soon after the explosion. We find that the circumstellar medium around red supergiants (especially massive ones) and the circumstellar medium dense enough to make Type IIn supernovae can establish collisional ionization equilibrium soon after the explosion and can evolve to become recombining supernova remnants. Wolf-Rayet stars and white dwarfs have the possibility to be recombining supernova remnants but the fraction is expected to be very small. As the occurrence rate of the explosions of red supergiants is much higher than that of Type IIn supernovae, the major progenitors of recombining supernova remnants are likely to be red supergiants.

  15. Human platelet lysate improves human cord blood derived ECFC survival and vasculogenesis in three dimensional (3D) collagen matrices.

    PubMed

    Kim, Hyojin; Prasain, Nutan; Vemula, Sasidhar; Ferkowicz, Michael J; Yoshimoto, Momoko; Voytik-Harbin, Sherry L; Yoder, Mervin C

    2015-09-01

    Human cord blood (CB) is enriched in circulating endothelial colony forming cells (ECFCs) that display high proliferative potential and in vivo vessel forming ability. Since diminished ECFC survival is known to dampen the vasculogenic response in vivo, we tested how long implanted ECFC survive and generate vessels in three-dimensional (3D) type I collagen matrices in vitro and in vivo. We hypothesized that human platelet lysate (HPL) would promote cell survival and enhance vasculogenesis in the 3D collagen matrices. We report that the percentage of ECFC co-cultured with HPL that were alive was significantly enhanced on days 1 and 3 post-matrix formation, compared to ECFC alone containing matrices. Also, co-culture of ECFC with HPL displayed significantly more vasculogenic activity compared to ECFC alone and expressed significantly more pro-survival molecules (pAkt, p-Bad and Bcl-xL) in the 3D collagen matrices in vitro. Treatment with Akt1 inhibitor (A-674563), Akt2 inhibitor (CCT128930) and Bcl-xL inhibitor (ABT-263/Navitoclax) significantly decreased the cell survival and vasculogenesis of ECFC co-cultured with or without HPL and implicated activation of the Akt1 pathway as the critical mediator of the HPL effect on ECFC in vitro. A significantly greater average vessel number and total vascular area of human CD31(+) vessels were present in implants containing ECFC and HPL, compared to the ECFC alone implants in vivo. We conclude that implantation of ECFC with HPL in vivo promotes vasculogenesis and augments blood vessel formation via diminishing apoptosis of the implanted ECFC. PMID:26122935

  16. Therapeutic Approach in the Improvement of Endothelial Dysfunction: The Current State of the Art

    PubMed Central

    Radenković, Miroslav; Stojanović, Marko; Potpara, Tatjana; Prostran, Milica

    2013-01-01

    The endothelium has a central role in the regulation of blood flow through continuous modulation of vascular tone. This is primarily accomplished by balanced release of endothelial relaxing and contractile factors. The healthy endothelial cells are essential for maintenance of vascular homeostasis involving antioxidant, anti-inflammatory, pro-fibrinolytic, anti-adhesive, or anticoagulant effects. Oppositely, endothelial dysfunction is primarily characterized by impaired regulation of vascular tone as a result of reduced endothelial nitric oxide (NO) synthase activity, lack of cofactors for NO synthesis, attenuated NO release, or increased NO degradation. So far, the pharmacological approach in improving/reversal of endothelial dysfunction was shown to be beneficial in clinical trials that have investigated actions of different cardiovascular drugs. The aim of this paper was to summarize some of the latest clinical findings related to therapeutic possibilities for improving endothelial dysfunction in different pathological conditions. In the majority of presented clinical investigations, the assessment of improvement or reversal of endothelial dysfunction was performed through the flow-mediated dilatation measurement, and in some of those endothelial progenitor cells' count was used for the same purpose. Still, given the fast and continuous development of this field, the evidence acquisition included the MEDLINE data base screening and the selection of articles published between 2010 and 2012. PMID:23509696

  17. A dynamic niche provides Kit ligand in a stage-specific manner to the earliest thymocyte progenitors

    PubMed Central

    Buono, Mario; Facchini, Raffaella; Matsuoka, Sahoko; Thongjuea, Supat; Waithe, Dominique; Luis, Tiago C.; Giustacchini, Alice; Besmer, Peter; Mead, Adam J.; Jacobsen, Sten Eirik W.; Nerlov, Claus

    2016-01-01

    Thymic T-cell development is initiated from bone marrow-derived multi-potent thymus seeding progenitors (TSPs). During the early stages of thymocyte differentiation progenitors become T-cell restricted. However, the cellular environments supporting these critical initial stages of T-cell development within the thymic cortex are not known. We here use the dependence of early, c-Kit–expressing thymic progenitors on Kit ligand (KitL) to show that CD4–CD8–c-Kit+CD25– DN1-stage progenitors associate with, and depend on the membrane-bound form of KitL (mKitL) provided by, a cortex-specific KitL-expressing vascular endothelial cell (VEC) population. In contrast, the subsequent CD4–CD8–c-Kit+CD25+ DN2 stage progenitors associate selectively with cortical thymic epithelial cells (cTECs) and depend on cTEC-presented mKitL. These results show that the dynamic process of early thymic progenitor differentiation is paralleled by migration-dependent changes to the supporting niche, and identify VECs as a thymic niche cell, with mKitL as a critical ligand. PMID:26780297

  18. The Involving Roles of Intrahepatic and Extrahepatic Stem/Progenitor Cells (SPCs) to Liver Regeneration.

    PubMed

    Liu, Wei-Hui; Ren, Li-Na; Wang, Tao; Navarro-Alvarez, Nalu; Tang, Li-Jun

    2016-01-01

    Liver regeneration is usually attributed to mature hepatocytes, which possess a remarkable potential to proliferate under mild to moderate injury. However, when the liver is severely damaged or hepatocyte proliferation is greatly inhibited, liver stem/progenitor cells (LSPCs) will contribute to the liver regeneration process. LSPCs in the developing liver have been extensively characterized, however, their contributing role to liver regeneration has not been completely understood. In addition to the restoration of the liver parenchymal tissue by hepatocytes or/and LSPCs, or in some cases bone marrow (BM) derived cells, such as hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs), the wound healing after injury in terms of angiopoiesis by liver sinusoidal endothelial cells (LSECs) or/and sinusoidal endothelial progenitor cells (SEPCs) is another important aspect taking place during regeneration. To conclude, liver regeneration can be mainly divided into three distinct restoring levels according to the cause and severity of injury: hepatocyte dominant regeneration, LSPCs mediated regeneration, extrahepatic stem cells participative regeneration. In this review, we focus on the recent findings of liver regeneration, especially on those related to stem/progenitor cells (SPCs)-mediated regeneration and their potential clinical applications and challenges. PMID:27489499

  19. The Involving Roles of Intrahepatic and Extrahepatic Stem/Progenitor Cells (SPCs) to Liver Regeneration

    PubMed Central

    Liu, Wei-hui; Ren, Li-na; Wang, Tao; Navarro-Alvarez, Nalu; Tang, Li-jun

    2016-01-01

    Liver regeneration is usually attributed to mature hepatocytes, which possess a remarkable potential to proliferate under mild to moderate injury. However, when the liver is severely damaged or hepatocyte proliferation is greatly inhibited, liver stem/progenitor cells (LSPCs) will contribute to the liver regeneration process. LSPCs in the developing liver have been extensively characterized, however, their contributing role to liver regeneration has not been completely understood. In addition to the restoration of the liver parenchymal tissue by hepatocytes or/and LSPCs, or in some cases bone marrow (BM) derived cells, such as hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs), the wound healing after injury in terms of angiopoiesis by liver sinusoidal endothelial cells (LSECs) or/and sinusoidal endothelial progenitor cells (SEPCs) is another important aspect taking place during regeneration. To conclude, liver regeneration can be mainly divided into three distinct restoring levels according to the cause and severity of injury: hepatocyte dominant regeneration, LSPCs mediated regeneration, extrahepatic stem cells participative regeneration. In this review, we focus on the recent findings of liver regeneration, especially on those related to stem/progenitor cells (SPCs)-mediated regeneration and their potential clinical applications and challenges. PMID:27489499

  20. EA-1, a novel adhesion molecule involved in the homing of progenitor T lymphocytes to the thymus

    PubMed Central

    1991-01-01

    The mouse progenitor T lymphocyte (pro-T) cell line FTF1 binds in vitro to thymus blood vessels, the thymic capsule, and liver from newborn mice. A mAb, EA-1, raised against an embryonic mouse endothelial cell line, blocked adhesion. The antibody also interfered with pro-T cell adhesion to a thymus-derived mouse endothelial cell line; it had no effect on the adhesion of mature T lymphocytes and myeloid cells. The antigen recognized by EA-1 is located on the vascular endothelium of various mouse tissues and absent on pro-T cells. EA-1 antibody precipitates molecules with apparent molecular weights of 110,000, 140,000, 160,000, and 200,000. Immunoclearing and binding-inhibition studies with antibodies against known adhesion molecules suggest that the EA-1 antigen is a novel adhesion molecule involved in colonization of the embryonic thymus by T cell progenitors. PMID:1874787

  1. Angiocrine functions of organ-specific endothelial cells

    PubMed Central

    Rafii, Shahin; Butler, Jason M; Ding, Bi-Sen

    2016-01-01

    Preface Endothelial cells lining blood vessel capillaries are not just passive conduits for delivering blood. Tissue-specific endothelium establish specialized vascular niches that deploy specific sets of growth factors, known as angiocrine factors, which actively participate in inducing, specifying, patterning, and guiding organ regeneration and maintaining homeostasis and metabolism. Angiocrine factors upregulated in response to injury orchestrates self-renewal and differentiation of tissue-specific repopulating resident stem and progenitor cells into functional organs. Uncovering the precise mechanisms whereby physiological-levels of angiocrine factors are spatially and temporally produced, and distributed by organotypic endothelium to repopulating cells, will lay the foundation for driving organ repair without scarring. PMID:26791722

  2. Modeling renal progenitors - defining the niche.

    PubMed

    Tanigawa, Shunsuke; Perantoni, Alan O

    2016-01-01

    Significant recent advances in methodologies for the differentiation of pluripotent stem cells to renal progenitors as well as the definition of niche conditions for sustaining those progenitors have dramatically enhanced our understanding of their biology and developmental programing, prerequisites for establishing viable approaches to renal regeneration. In this article, we review the evolution of culture techniques and models for the study of metanephric development, describe the signaling mechanisms likely to be driving progenitor self-renewal, and discuss current efforts to generate de novo functional tissues, providing in depth protocols and niche conditions for the stabilization of the nephronic Six2+progenitor. PMID:26856661

  3. Evolution of endothelial keratoplasty.

    PubMed

    Price, Francis W; Price, Marianne O

    2013-11-01

    Endothelial keratoplasty has evolved into a popular alternative to penetrating keratoplasty (PK) for the treatment of endothelial dysfunction. Although the earliest iterations were challenging and were not widely adopted, the iteration known as Descemet stripping endothelial keratoplasty (DSEK) has gained widespread acceptance. DSEK combines a simplified technique for stripping dysfunctional endothelium from the host cornea and microkeratome dissection of the donor tissue, a step now commonly completed in advance by eye bank technicians. Studies show that a newer endothelial keratoplasty iteration, known as Descemet membrane endothelial keratoplasty (DMEK), provides an even faster and better visual recovery than DSEK does. In addition, DMEK significantly reduces the risk of immunologic graft rejection episodes compared with that in DSEK or in PK. Although the DMEK donor tissue, consisting of the bare endothelium and Descemet membrane without any stroma, is more challenging to prepare and position in the recipient eye, recent improvements in instrumentation and surgical techniques are increasing the ease and the reliability of the procedure. DSEK successfully mitigates 2 of the main liabilities of PK: ocular surface complications and structural problems (including induced astigmatism and perpetually weak wounds), whereas DMEK further mitigates the 2 principal remaining liabilities of PK: immunologic graft reactions and secondary glaucoma from prolonged topical corticosteroid use.

  4. Frs2α-deficiency in cardiac progenitors disrupts a subset of FGF signals required for outflow tract morphogenesis

    PubMed Central

    Zhang, Jue; Lin, Yongshun; Zhang, Yongyou; Lan, Yongsheng; Lin, Chunhong; Moon, Anne M.; Schwartz, Robert J.; Martin, James F.; Wang, Fen

    2009-01-01

    Summary The cardiac outflow tract (OFT) is a developmentally complex structure derived from multiple lineages and is often defective in human congenital anomalies. While emerging evidence shows that the fibroblast growth factor (FGF) is essential for OFT development, the downstream pathways mediating FGF-signaling in cardiac progenitors remain poorly understood. Here, we report that FRS2α, an adaptor protein that links FGF receptor kinases to multiple signaling pathways, mediates critical aspects of FGF-dependent OFT development. Ablation of Frs2α in mesodermal OFT progenitor cells that originate in the second heart field (SHF) affects their expansion into the OFT myocardium, resulting in OFT misalignment and hypoplasia. Moreover, Frs2α mutants had defective endothelial-mesenchymal-transition and neural crest cell recruitment into the OFT cushions, resulting in OFT septation defects. The results provide new insight into the signaling molecules downstream of FGF receptor tyrosine kinases in cardiac progenitors. PMID:18832393

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

    PubMed

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

    2013-09-01

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

  6. Induction of hematopoietic and endothelial cell program orchestrated by ETS transcription factor ER71/ETV2

    PubMed Central

    Liu, Fang; Li, Daofeng; Yu, Yik Yeung Lawrence; Kang, Inyoung; Cha, Min-Ji; Kim, Ju Young; Park, Changwon; Watson, Dennis K; Wang, Ting; Choi, Kyunghee

    2015-01-01

    The ETS factor ETV2 (aka ER71) is essential for the generation of the blood and vascular system, as ETV2 deficiency leads to a complete block in blood and endothelial cell formation and embryonic lethality in the mouse. However, the ETV2-mediated gene regulatory network and signaling governing hematopoietic and endothelial cell development are poorly understood. Here, we map ETV2 global binding sites and carry out in vitro differentiation of embryonic stem cells, and germ line and conditional knockout mouse studies to uncover mechanisms involved in the hemangiogenic fate commitment from mesoderm. We show that ETV2 binds to enhancers that specify hematopoietic and endothelial cell lineages. We find that the hemangiogenic progenitor population in the developing embryo can be identified as FLK1highPDGFRα−. Notably, these hemangiogenic progenitors are exclusively sensitive to ETV2-dependent FLK1 signaling. Importantly, ETV2 turns on other Ets genes, thereby establishing an ETS hierarchy. Consequently, the hematopoietic and endothelial cell program initiated by ETV2 is maintained partly by other ETS factors through an ETS switching mechanism. These findings highlight the critical role that transient ETV2 expression plays in the regulation of hematopoietic and endothelial cell lineage specification and stability. PMID:25802403

  7. Induction of hematopoietic and endothelial cell program orchestrated by ETS transcription factor ER71/ETV2.

    PubMed

    Liu, Fang; Li, Daofeng; Yu, Yik Yeung Lawrence; Kang, Inyoung; Cha, Min-Ji; Kim, Ju Young; Park, Changwon; Watson, Dennis K; Wang, Ting; Choi, Kyunghee

    2015-05-01

    The ETS factor ETV2 (aka ER71) is essential for the generation of the blood and vascular system, as ETV2 deficiency leads to a complete block in blood and endothelial cell formation and embryonic lethality in the mouse. However, the ETV2-mediated gene regulatory network and signaling governing hematopoietic and endothelial cell development are poorly understood. Here, we map ETV2 global binding sites and carry out in vitro differentiation of embryonic stem cells, and germ line and conditional knockout mouse studies to uncover mechanisms involved in the hemangiogenic fate commitment from mesoderm. We show that ETV2 binds to enhancers that specify hematopoietic and endothelial cell lineages. We find that the hemangiogenic progenitor population in the developing embryo can be identified as FLK1(high)PDGFRα(-). Notably, these hemangiogenic progenitors are exclusively sensitive to ETV2-dependent FLK1 signaling. Importantly, ETV2 turns on other Ets genes, thereby establishing an ETS hierarchy. Consequently, the hematopoietic and endothelial cell program initiated by ETV2 is maintained partly by other ETS factors through an ETS switching mechanism. These findings highlight the critical role that transient ETV2 expression plays in the regulation of hematopoietic and endothelial cell lineage specification and stability.

  8. Critical Roles of Reactive Oxygen Species in Age-Related Impairment in Ischemia-Induced Neovascularization by Regulating Stem and Progenitor Cell Function

    PubMed Central

    Lam, Yuen Ting

    2016-01-01

    Reactive oxygen species (ROS) regulate bone marrow microenvironment for stem and progenitor cells functions including self-renewal, differentiation, and cell senescence. In response to ischemia, ROS also play a critical role in mediating the mobilization of endothelial progenitor cells (EPCs) from the bone marrow to the sites of ischemic injury, which contributes to postnatal neovascularization. Aging is an unavoidable biological deteriorative process with a progressive decline in physiological functions. It is associated with increased oxidative stress and impaired ischemia-induced neovascularization. This review discusses the roles of ROS in regulating stem and progenitor cell function, highlighting the impact of unbalanced ROS levels on EPC dysfunction and the association with age-related impairment in ischemia-induced neovascularization. Furthermore, it discusses strategies that modulate the oxidative levels of stem and progenitor cells to enhance the therapeutic potential for elderly patients with cardiovascular disease. PMID:26697140

  9. Isolation of neural crest derived chromaffin progenitors from adult adrenal medulla.

    PubMed

    Chung, Kuei-Fang; Sicard, Flavie; Vukicevic, Vladimir; Hermann, Andreas; Storch, Alexander; Huttner, Wieland B; Bornstein, Stefan R; Ehrhart-Bornstein, Monika

    2009-10-01

    Chromaffin cells of the adrenal medulla are neural crest-derived cells of the sympathoadrenal lineage. Unlike the closely-related sympathetic neurons, a subpopulation of proliferation-competent cells exists even in the adult. Here, we describe the isolation, expansion, and in vitro characterization of proliferation-competent progenitor cells from the bovine adrenal medulla. Similar to neurospheres, these cells, when prevented from adherence to the culture dish, grew in spheres, which we named chromospheres. These chromospheres were devoid of mRNA specific for smooth muscle cells (MYH11) or endothelial cells (PECAM1). During sphere formation, markers for differentiated chromaffin cells, such as phenylethanolamine-N-methyl transferase, were downregulated while neural progenitor markers nestin, vimentin, musashi 1, and nerve growth factor receptor, as well as markers of neural crest progenitor cells such as Sox1 and Sox9, were upregulated. Clonal analysis and bromo-2'-deoxyuridine-incorporation analysis demonstrated the self-renewing capacity of chromosphere cells. Differentiation protocols using NGF and BMP4 or dexamethasone induced neuronal or endocrine differentiation, respectively. Electrophysiological analyses of neural cells derived from chromospheres revealed functional properties of mature nerve cells, such as tetrodotoxin-sensitive sodium channels and action potentials. Our study provides evidence that proliferation and differentiation competent chromaffin progenitor cells can be isolated from adult adrenal medulla and that these cells might harbor the potential for the treatment of neurodegenerative diseases, such as Parkinson's disease. PMID:19609938

  10. Hepatocyte growth factor induces proliferation and differentiation of multipotent and erythroid hemopoietic progenitors

    PubMed Central

    1994-01-01

    Hepatocyte growth factor (HGF) is a mesenchymal derived growth factor known to induce proliferation and "scattering" of epithelial and endothelial cells. Its receptor is the tyrosine kinase encoded by the c- MET protooncogene. Here we show that highly purified recombinant HGF stimulates hemopoietic progenitors to form colonies in vitro. In the presence of erythropoietin, picomolar concentrations of HGF induced the formation of erythroid burst-forming unit colonies from CD34-positive cells purified from human bone marrow, peripheral blood, or umbilical cord blood. The growth stimulatory activity was restricted to the erythroid lineage. HGF also stimulated the formation of multipotent CFU- GEMM colonies. This effect is synergized by stem cell factor, the ligand of the tyrosine kinase receptor encoded by the c-KIT protooncogene, which is active on early hemopoietic progenitors. By flow cytometry analysis, the receptor for HGF was found to be expressed on the cell surface in a fraction of CD34+ progenitors. Moreover, in situ hybridization experiments showed that HGF receptor mRNA is highly expressed in embryonic erythroid cells (megaloblasts). HGF mRNA was also found to be produced in the embryonal liver. These data show that HGF plays a direct role in the control of proliferation and differentiation of erythroid progenitors, and they suggest that it may be one of the long-sought mediators of paracrine interactions between stromal and hemopoietic cells within the hemopoietic microenvironment. PMID:7528222

  11. [Bone and Stem Cells. Bone marrow microenvironment niches for hematopoietic stem and progenitor cells].

    PubMed

    Nagasawa, Takashi

    2014-04-01

    In bone marrow, the special microenvironments known as niches control proliferation and differentiation of hematopoietic stem and progenitor cells (HSPCs) . However, the identity and functions of the niches has been a subject of longstanding debate. Although it has been reported previously that osteoblasts lining the bone surface act as HSC niches, their precise role in HSC maintenance remains unclear. On the other hand, the adipo-osteogenic progenitors with long processes, termed CXCL12-abundant reticular (CAR) cells, which preferentially express the chemokine CXCL12, stem cell factor (SCF) , leptin receptor and PDGF receptor-β were identified in the bone marrow. Recent studies revealed that endothelial cells of bone marrow vascular sinuses and CAR cells provided niches for HSCs. The identity and functions of various other candidate HSC niche cells, including nestin-expressing cells and Schwann cells would also be discussed in this review.

  12. Dysregulation of VEGF-induced proangiogenic Ca2+ oscillations in primary myelofibrosis-derived endothelial colony-forming cells.

    PubMed

    Dragoni, Silvia; Reforgiato, Marta; Zuccolo, Estella; Poletto, Valentina; Lodola, Francesco; Ruffinatti, Federico Alessandro; Bonetti, Elisa; Guerra, Germano; Barosi, Giovanni; Rosti, Vittorio; Moccia, Francesco

    2015-12-01

    Endothelial progenitor cells could be implicated in the aberrant neoangiogenesis that occurs in bone marrow and spleen in patients with primary myelofibrosis (PMF). However, antivascular endothelial growth factor (VEGF) monotherapy had only a modest and transient effect in these individuals. Recently it was found that VEGF-induced proangiogenic intracellular Ca(2+) oscillations could be impaired in endothelial progenitor cells of subjects with malignancies. Therefore, we employed Ca(2+) imaging, wavelet analysis, and functional assays to assess whether and how VEGF-induced Ca(2+) oscillations are altered in PMF-derived endothelial progenitor cells. We focused on endothelial colony-forming cells (ECFCs), which are the only endothelial progenitor cell subtype capable of forming neovessels both in vivo and in vitro. VEGF triggers repetitive Ca(2+) spikes in both normal ECFCs (N-ECFCs) and ECFCs obtained from PMF patients (PMF-ECFCs). However, the spiking response to VEGF is significantly weaker in PMF-ECFCs. VEGF-elicited Ca(2+) oscillations are patterned by the interaction between inositol-1,4,5-trisphosphate-dependent Ca(2+) mobilization and store-operated Ca(2+) entry. However, in most PMF-ECFCs, Ca(2+) oscillations are triggered by a store-independent Ca(2+) entry pathway. We found that diacylglycerol gates transient receptor potential canonical 1 channel to trigger VEGF-dependent Ca(2+) spikes by recruiting the phospholipase C/inositol-1,4,5-trisphosphate signaling pathway, reflected as a decrease in endoplasmic reticulum Ca(2+) content. Finally, we found that, apart from being less robust and dysregulated as compared with N-ECFCs, VEGF-induced Ca(2+) oscillations modestly stimulate PMF-ECFC growth and in vitro angiogenesis. These results may explain the modest effect of anti-VEGF therapies in PMF. PMID:26432919

  13. Mechanisms of Tubulogenesis and Endothelial Phenotype Expression by MSCs

    PubMed Central

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

    2015-01-01

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

  14. Is exercise training an effective therapy targeting endothelial dysfunction and vascular wall inflammation?

    PubMed

    Ribeiro, Fernando; Alves, Alberto Jorge; Duarte, José Alberto; Oliveira, José

    2010-06-11

    There is an increasing evidence that endothelial dysfunction and vascular wall inflammation are present in all stages of atherosclerosis. Atherosclerosis does not have to necessarily progress to an acute clinical event. Several therapeutic strategies exist, such as exercise training, which mitigates endothelial dysfunction and inflammation. Exercise training consistently improves the nitric oxide bioavailability, and the number of endothelial progenitor cells, and also diminishes the level of inflammatory markers, namely pro-inflammatory cytokines and C-reactive protein. However, the mechanisms by which exercise improves endothelial function in coronary artery disease patients are not fully clarified. Several mechanisms have been proposed to explain the positive effect of exercise on the disease progression. They include the decrease in cytokine production by the adipose tissue, skeletal muscles, endothelial cells, and blood mononuclear cells, and also, the increase in the bioavailability of nitric oxide, antioxidant defences, and regenerative capacity of endothelium. This study aims to provide a critical review of the literature linking exercise, inflammation, and endothelial dysfunction in coronary artery patients, and to discuss the potential mechanisms behind the exercise-training improvement of endothelial function and inflammatory status.

  15. Autophagy Proteins ATG5 and ATG7 Are Essential for the Maintenance of Human CD34(+) Hematopoietic Stem-Progenitor Cells.

    PubMed

    Gomez-Puerto, Maria Catalina; Folkerts, Hendrik; Wierenga, Albertus T J; Schepers, Koen; Schuringa, Jan Jacob; Coffer, Paul J; Vellenga, Edo

    2016-06-01

    Autophagy is a highly regulated catabolic process that involves sequestration and lysosomal degradation of cytosolic components such as damaged organelles and misfolded proteins. While autophagy can be considered to be a general cellular housekeeping process, it has become clear that it may also play cell type-dependent functional roles. In this study, we analyzed the functional importance of autophagy in human hematopoietic stem/progenitor cells (HSPCs), and how this is regulated during differentiation. Western blot-based analysis of LC3-II and p62 levels, as well as flow cytometry-based autophagic vesicle quantification, demonstrated that umbilical cord blood-derived CD34(+) /CD38(-) immature hematopoietic progenitors show a higher autophagic flux than CD34(+) /CD38(+) progenitors and more differentiated myeloid and erythroid cells. This high autophagic flux was critical for maintaining stem and progenitor function since knockdown of autophagy genes ATG5 or ATG7 resulted in reduced HSPC frequencies in vitro as well as in vivo. The reduction in HSPCs was not due to impaired differentiation, but at least in part due to reduced cell cycle progression and increased apoptosis. This is accompanied by increased expression of p53, proapoptotic genes BAX and PUMA, and the cell cycle inhibitor p21, as well as increased levels of cleaved caspase-3 and reactive oxygen species. Taken together, our data demonstrate that autophagy is an important regulatory mechanism for human HSCs and their progeny, reducing cellular stress and promoting survival. Stem Cells 2016;34:1651-1663.

  16. Biophysical Properties of Scaffolds Modulate Human Blood Vessel Formation from Circulating Endothelial Colony-Forming Cells

    NASA Astrophysics Data System (ADS)

    Critser, Paul J.; Yoder, Mervin C.

    A functional vascular system forms early in development and is continually remodeled throughout the life of the organism. Impairment to the regeneration or repair of this system leads to tissue ischemia, dysfunction, and disease. The process of vascular formation and remodeling is complex, relying on local microenvironmental cues, cytokine signaling, and multiple cell types to function properly. Tissue engineering strategies have attempted to exploit these mechanisms to develop functional vascular networks for the generation of artificial tissues and therapeutic strategies to restore tissue homeostasis. The success of these strategies requires the isolation of appropriate progenitor cell sources which are straightforward to obtain, display high proliferative potential, and demonstrate an ability to form functional vessels. Several populations are of interest including endothelial colony-forming cells, a subpopulation of endothelial progenitor cells. Additionally, the development of scaffolds to deliver and support progenitor cell survival and function is crucial for the formation of functional vascular networks. The composition and biophysical properties of these scaffolds have been shown to modulate endothelial cell behavior and vessel formation. However, further investigation is needed to better understand how these mechanical properties and biophysical properties impact vessel formation. Additionally, several other cell populations are involved in neoangiogenesis and formation of tissue parenchyma and an understanding of the potential impact of these cell populations on the biophysical properties of scaffolds will also be needed to advance these strategies. This chapter examines how the biophysical properties of matrix scaffolds can influence vessel formation and remodeling and, in particular, the impact on in vivo human endothelial progenitor cell vessel formation.

  17. Preterm Cord Blood Contains a Higher Proportion of Immature Hematopoietic Progenitors Compared to Term Samples

    PubMed Central

    Podestà, Marina; Bruschettini, Matteo; Cossu, Claudia; Sabatini, Federica; Dagnino, Monica; Romantsik, Olga; Spaggiari, Grazia Maria; Ramenghi, Luca Antonio; Frassoni, Francesco

    2015-01-01

    Background Cord blood contains high number of hematopoietic cells that after birth disappear. In this paper we have studied the functional properties of the umbilical cord blood progenitor cells collected from term and preterm neonates to establish whether quantitative and/or qualitative differences exist between the two groups. Methods and Results Our results indicate that the percentage of total CD34+ cells was significantly higher in preterm infants compared to full term: 0.61% (range 0.15–4.8) vs 0.3% (0.032–2.23) p = 0.0001 and in neonates <32 weeks of gestational age (GA) compared to those ≥32 wks GA: 0.95% (range 0.18–4.8) and 0.36% (0.15–3.2) respectively p = 0.0025. The majority of CD34+ cells co-expressed CD71 antigen (p<0.05 preterm vs term) and grew in vitro large BFU-E, mostly in the second generation. The subpopulations CD34+CD38- and CD34+CD45- resulted more represented in preterm samples compared to term, conversely, Side Population (SP) did not show any difference between the two group. The absolute number of preterm colonies (CFCs/10microL) resulted higher compared to term (p = 0.004) and these progenitors were able to grow until the third generation maintaining an higher proportion of CD34+ cells (p = 0.0017). The number of colony also inversely correlated with the gestational age (Pearson r = -0.3001 p<0.0168). Conclusions We found no differences in the isolation and expansion capacity of Endothelial Colony Forming Cells (ECFCs) from cord blood of term and preterm neonates: both groups grew in vitro large number of endothelial cells until the third generation and showed a transitional phenotype between mesenchymal stem cells and endothelial progenitors (CD73, CD31, CD34 and CD144)The presence, in the cord blood of preterm babies, of high number of immature hematopoietic progenitors and endothelial/mesenchymal stem cells with high proliferative potential makes this tissue an important source of cells for developing new cells therapies

  18. Progenitors of type Ia supernovae

    NASA Astrophysics Data System (ADS)

    Raskin, Cody

    Type Ia supernovae are important, but mysterious cosmological tools. Their standard brightnesses have enabled cosmologists to measure extreme distances and to discover dark energy. However, the nature of their progenitor mechanisms remains elusive, with many competing models offering only partial clues to their origins. Here, type Ia supernova delay times are explored using analytical models. Combined with a new observation technique, this model places new constraints on the characteristic time delay between the formation of stars and the first type Ia supernovae. This derived delay time (500 million years) implies low-mass companions for single degenerate progenitor scenarios. In the latter portions of this dissertation, two progenitor mechanisms are simulated in detail; white dwarf collisions and mergers. From the first of these simulations, it is evident that white dwarf collisions offer a viable and unique pathway to producing type Ia supernovae. Many of the combinations of masses simulated produce sufficient quantities of 56 Ni (up to 0.51 solar masses) to masquerade as normal type Ia supernovae. Other combinations of masses produce 56 Ni yields that span the entire range of supernova brightnesses, from the very dim and underluminous, with 0.14 solar masses, to the over-bright and superluminous, with up to 1.71 solar masses. The 56 Ni yield in the collision simulations depends non-linearly on total system mass, mass ratio, and impact parameter. Using the same numerical tools as in the collisions examination, white dwarf mergers are studied in detail. Nearly all of the simulations produce merger remnants consisting of a cold, degenerate core surrounded by a hot accretion disk. The properties of these disks have strong implications for various viscosity treatments that have attempted to pin down the accretion times. Some mass combinations produce super-Chandrasekhar cores on shorter time scales than viscosity driven accretion. A handful of simulations also

  19. A boost of BMP4 accelerates the commitment of human embryonic stem cells to the endothelial lineage.

    PubMed

    Goldman, Orit; Feraud, Olivier; Boyer-Di Ponio, Julie; Driancourt, Catherine; Clay, Denis; Le Bousse-Kerdiles, Marie-Caroline; Bennaceur-Griscelli, Annelise; Uzan, Georges

    2009-08-01

    Embryoid bodies (EBs) generated during differentiation of human embryonic stem cells (hESCs) contain vascular-like structures, suggesting that commitment of mesoderm progenitors into endothelial cells occurs spontaneously. We showed that bone morphogenetic protein 4 (BMP4), an inducer of mesoderm, accelerates the peak expression of CD133/kinase insert domain-containing receptor (KDR) and CD144/KDR. Because the CD133(+)KDR(+) population could represent endothelial progenitors, we sorted them at day 7 and cultured them in endothelial medium. These cells were, however, unable to differentiate into endothelial cells. Under standard conditions, the CD144(+)KDR(+) population represents up to 10% of the total cells at day 12. In culture, these cells, if sorted, give rise to a homogeneous population with a morphology typical of endothelial cells and express endothelial markers. These endothelial cells derived from the day 12 sorted population were functional, as assessed by different in vitro assays. When EBs were stimulated by BMP4, the CD144(+)KDR(+) peak was shifted to day 7. Most of these cells, however, were CD31(-), becoming CD31(+) in culture. They then expressed von Willebrand factor and were functional. This suggests that, initially, the BMP4-boosted day 7, CD144(+)KDR(+)CD31(-) population represents immature endothelial cells that differentiate into mature endothelial cells in culture. The expression of OCT3/4, a marker of immaturity for hESCs decreases during EB differentiation, decreasing faster following BMP4 induction. We also show that BMP4 inhibits the global expression of GATA2 and RUNX1, two transcription factors involved in hemangioblast formation, at day 7 and day 12.

  20. Prorenin receptor is critical for nephron progenitors.

    PubMed

    Song, Renfang; Preston, Graeme; Kidd, Laura; Bushnell, Daniel; Sims-Lucas, Sunder; Bates, Carlton M; Yosypiv, Ihor V

    2016-01-15

    Deficient nephrogenesis is the major factor contributing to renal hypoplasia defined as abnormally small kidneys. Nephron induction during kidney development is driven by reciprocal interactions between progenitor cells of the cap mesenchyme (CM) and the ureteric bud (UB). The prorenin receptor (PRR) is a receptor for renin and prorenin, and an accessory subunit of the vacuolar proton pump H(+)-ATPase. Global loss of PRR is lethal in mice and PRR mutations are associated with a high blood pressure, left ventricular hypertrophy and X-linked mental retardation in humans. To circumvent lethality of the ubiquitous PRR mutation in mice and to determine the potential role of the PRR in nephrogenesis, we generated a mouse model with a conditional deletion of the PRR in Six2(+) nephron progenitors and their epithelial derivatives (Six2(PRR-/-)). Targeted ablation of PRR in Six2(+) nephron progenitors caused a marked decrease in the number of developing nephrons, small cystic kidneys and podocyte foot process effacement at birth, and early postnatal death. Reduced congenital nephron endowment resulted from premature depletion of nephron progenitor cell population due to impaired progenitor cell proliferation and loss of normal molecular inductive response to canonical Wnt/β-catenin signaling within the metanephric mesenchyme. At 2 months of age, heterozygous Six2(PRR+/-) mice exhibited focal glomerulosclerosis, decreased kidney function and massive proteinuria. Collectively, these findings demonstrate a cell-autonomous requirement for the PRR within nephron progenitors for progenitor maintenance, progression of nephrogenesis, normal kidney development and function.

  1. Accumulation of Multipotent Progenitor Cells on Polymethylpentene Membranes During Extracorporeal Membrane Oxygenation.

    PubMed

    Lehle, Karla; Friedl, Lucas; Wilm, Julius; Philipp, Alois; Müller, Thomas; Lubnow, Matthias; Schmid, Christof

    2016-06-01

    Multipotent progenitor cells were mobilized during pediatric extracorporeal membrane oxygenation (ECMO). We hypothesize that these cells also adhered onto polymethylpentene (PMP) fibers within the membrane oxygenator (MO) during adult ECMO support. Mononuclear cells were removed from the surface of explanted PMP-MOs (n = 16). Endothelial-like outgrowth and mesenchymal-like cells were characterized by flow cytometric analysis using different surface markers. Spindle-shaped attaching cells were identified early, but without proliferative activity. After long-term cultivation palisading type or cobblestone-type outgrowth cells with high proliferative activity appeared and were characterized as (i) leukocytoid CD45+/CD31+ (CD133+/VEGFR-II+/CD90+/CD14+/CD146dim/CD105dim); (ii) endothelial-like CD45-/CD31+ (VEGF-RII+/CD146+/CD105+/CD133-/CD14-/CD90-); and (iii) mesenchymal-like cells CD45-/CD31- (CD105+/CD90+/CD133dim/VEGFR-II-/CD146-/CD14-). The distribution of the cell populations depended on the MO and cultivation time. Endothelial-like cells formed capillary-like structures and did uptake Dil-acetylated low-density lipoprotein. Endothelial- and mesenchymal-like cells adhered on the surface of PMP-MOs. Further research is needed to identify the clinical relevance of these cells.

  2. Progenitor's Signatures in Type Ia Supernova Remnants

    NASA Astrophysics Data System (ADS)

    Chiotellis, A.; Kosenko, D.; Schure, K. M.; Vink, J.

    2013-01-01

    The remnants of Type Ia supernovae (SNe Ia) can provide important clues about their progenitor histories. We discuss two well-observed supernova remnants (SNRs) that are believed to have resulted from SNe Ia, and use various tools to shed light on the possible progenitor histories. We find that Kepler's SNR is consistent with a symbiotic binary progenitor consisting of a white dwarf and an AGB star. Our hydrosimulations can reproduce the observed kinematic and morphological properties. For Tycho's remnant we use the characteristics of the X-ray spectrum and kinematics to show that the ejecta has likely interacted with dense circumstellar gas.

  3. Progenitor genealogy in the developing cerebral cortex.

    PubMed

    Laguesse, Sophie; Peyre, Elise; Nguyen, Laurent

    2015-01-01

    The mammalian cerebral cortex is characterized by a complex histological organization that reflects the spatio-temporal stratifications of related stem and neural progenitor cells, which are responsible for the generation of distinct glial and neuronal subtypes during development. Some work has been done to shed light on the existing filiations between these progenitors as well as their respective contribution to cortical neurogenesis. The aim of the present review is to summarize the current views of progenitor hierarchy and relationship in the developing cortex and to further discuss future research directions that would help us to understand the molecular and cellular regulating mechanisms involved in cerebral corticogenesis. PMID:25141969

  4. Fibroblast-Endothelial Partners for Vascularization Strategies in Tissue Engineering

    PubMed Central

    Costa-Almeida, Raquel; Gomez-Lazaro, Maria; Ramalho, Carla; Soares, Raquel; Guerreiro, Susana G.

    2015-01-01

    Cell-based approaches have emerged as a promising therapy to achieve successful vascularization in tissue engineering. Since fibroblasts activation and migration is required for physiological events relying on angiogenesis, we hypothesize herein that different fibroblasts exhibit distinct capacity to promote capillary-like structures assembly, by mature and progenitor endothelial cells (ECs). Outgrowth endothelial cells (OECs) were isolated from human umbilical cord blood samples and characterized by immunofluorescence and imaging flow cytometry for endothelial markers. Coculture systems were established using either human umbilical vein ECs (HUVECs) or OECs with fibroblasts, being evaluated at 7, 14, and 21 days of culture. Two types of human dermal fibroblasts (HDF) were used, namely neonatal human foreskin fibroblasts-1 (HFF-1) and juvenile HDF. OECs expressed EC markers and formed capillary-like structures. HFF-1 exhibited higher expression of transglutaminase-2, while HDF exhibited a higher expression of α-smooth muscle actin (α-SMA) and podoplanin, which were not observed for HFF-1. Formation of capillary-like structures was only observed in cocultures with HDF and not with HFF-1. No significant differences were found between HDF and OECs or HUVECs cocultures. These findings suggest that HDF is a preferential cell source for promoting vascularization, either using mature or progenitor ECs, probably due to their higher expression of α-SMA and podoplanin, and increased synthesis of extracellular matrix. This work opens new research possibilities regarding the use of specific fibroblast populations cocultured with ECs, as efficient partners for vascular development in regenerative medicine strategies. PMID:25340984

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

    PubMed Central

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

    2016-01-01

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

  6. In vitro differentiation of porcine aortic vascular precursor cells to endothelial and vascular smooth muscle cells.

    PubMed

    Zaniboni, Andrea; Bernardini, Chiara; Bertocchi, Martina; Zannoni, Augusta; Bianchi, Francesca; Avallone, Giancarlo; Mangano, Chiara; Sarli, Giuseppe; Calzà, Laura; Bacci, Maria Laura; Forni, Monica

    2015-09-01

    Recent findings suggest that progenitor and multipotent mesenchymal stromal cells (MSCs) are associated with vascular niches. Cells displaying mesenchymal properties and differentiating to whole components of a functional blood vessel, including endothelial and smooth muscle cells, can be defined as vascular stem cells (VSCs). Recently, we isolated a population of porcine aortic vascular precursor cells (pAVPCs), which have MSC- and pericyte-like properties. The aim of the present work was to investigate whether pAVPCs possess VSC-like properties and assess their differentiation potential toward endothelial and smooth muscle lineages. pAVPCs, maintained in a specific pericyte growth medium, were cultured in high-glucose DMEM + 10% FBS (long-term medium, LTM) or in human endothelial serum-free medium + 5% FBS and 50 ng/ml of hVEGF (endothelial differentiation medium, EDM). After 21 days of culture in LTM, pAVPCs showed an elongated fibroblast-like morphology, and they seem to organize in cord-like structures. qPCR analysis of smooth muscle markers [α-smooth muscle actin (α-SMA), calponin, and smooth muscle myosin (SMM) heavy chain] showed a significant increment of the transcripts, and immunofluorescence analysis confirmed the presence of α-SMA and SMM proteins. After 21 days of culture in EDM, pAVPCs displayed an endothelial cell-like morphology and revealed the upregulation of the expression of endothelial markers (CD31, vascular endothelial-cadherin, von Willebrand factor, and endothelial nitric oxide synthase) showing the CD31-typical pattern. In conclusion, pAVPCs could be defined as a VSC-like population considering that, if they are maintained in a specific pericyte medium, they express MSC markers, and they have, in addition to the classical mesenchymal trilineage differentiation potential, the capacity to differentiate in vitro toward the smooth muscle and the endothelial cell phenotypes.

  7. Endothelial PDGF-CC regulates angiogenesis-dependent thermogenesis in beige fat

    PubMed Central

    Seki, Takahiro; Hosaka, Kayoko; Lim, Sharon; Fischer, Carina; Honek, Jennifer; Yang, Yunlong; Andersson, Patrik; Nakamura, Masaki; Näslund, Erik; Ylä-Herttuala, Seppo; Sun, Meili; Iwamoto, Hideki; Li, Xuri; Liu, Yizhi; Samani, Nilesh J.; Cao, Yihai

    2016-01-01

    Cold- and β3-adrenoceptor agonist-induced sympathetic activation leads to angiogenesis and UCP1-dependent thermogenesis in mouse brown and white adipose tissues. Here we show that endothelial production of PDGF-CC during white adipose tissue (WAT) angiogenesis regulates WAT browning. We find that genetic deletion of endothelial VEGFR2, knockout of the Pdgf-c gene or pharmacological blockade of PDGFR-α impair the WAT-beige transition. We further show that PDGF-CC stimulation upregulates UCP1 expression and acquisition of a beige phenotype in differentiated mouse WAT-PDGFR-α+ progenitor cells, as well as in human WAT-PDGFR-α+ adipocytes, supporting the physiological relevance of our findings. Our data reveal a paracrine mechanism by which angiogenic endothelial cells modulate adipocyte metabolism, which may provide new targets for the treatment of obesity and related metabolic diseases. PMID:27492130

  8. Endothelialization of implanted cardiovascular biomaterial surfaces: the development from in vitro to in vivo.

    PubMed

    Liu, Tao; Liu, Shihui; Zhang, Kun; Chen, Junying; Huang, Nan

    2014-10-01

    Restenosis and thrombosis formation after cardiovascular devices implantation continue to be problematic. Although various platforms and parameters of cardiovascular devices have been designed and optimized over the years, postoperative complications are hard to avoid. The native vascular endothelium always provide a nonthrombogenic surface as well as prevent intimal overproliferation, thereby, the presence of a confluent endothelial cell layer on material surfaces have been widely accepted as an ideal approach to improve the biocompatibility of implanted cardiovascular materials. Endothelialization on biomaterial surfaces is initially developed by in vitro cell seeding. However, numerous no-perfect parts of this method are existed for clinical use. The emergency of endothelial progenitor cells may provide a promising way for setting these limitations. Over the last decades, countless researches about EPCs-based in vivo induced self-endothelialization have been reported and mainly focused on cellular therapy, pharmacological therapy, materials designing, or surface biofunctional modification. This review details the development of endothelialization on cardiovascular material surfaces from in vitro to in vivo. Endothelialization progress on the basis of molecular biological level and bioinformatics theory is expected to be the key point in the coming decades.

  9. Human progenitor cells derived from cardiac adipose tissue ameliorate myocardial infarction in rodents.

    PubMed

    Bayes-Genis, Antoni; Soler-Botija, Carolina; Farré, Jordi; Sepúlveda, Pilar; Raya, Angel; Roura, Santiago; Prat-Vidal, Cristina; Gálvez-Montón, Carolina; Montero, José Anastasio; Büscher, Dirk; Izpisúa Belmonte, Juan Carlos

    2010-11-01

    Myocardial infarction caused by vascular occlusion results in the formation of nonfunctional fibrous tissue. Cumulative evidence indicates that cell therapy modestly improves cardiac function; thus, novel cell sources with the potential to repair injured tissue are actively sought. Here, we identify and characterize a cell population of cardiac adipose tissue-derived progenitor cells (ATDPCs) from biopsies of human adult cardiac adipose tissue. Cardiac ATDPCs express a mesenchymal stem cell-like marker profile (strongly positive for CD105, CD44, CD166, CD29 and CD90) and have immunosuppressive capacity. Moreover, cardiac ATDPCs have an inherent cardiac-like phenotype and were able to express de novo myocardial and endothelial markers in vitro but not to differentiate into adipocytes. In addition, when cardiac ATDPCs were transplanted into injured myocardium in mouse and rat models of myocardial infarction, the engrafted cells expressed cardiac (troponin I, sarcomeric α-actinin) and endothelial (CD31) markers, vascularization increased, and infarct size was reduced in mice and rats. Moreover, significant differences between control and cell-treated groups were found in fractional shortening and ejection fraction, and the anterior wall remained significantly thicker 30days after cardiac delivery of ATDPCs. Finally, cardiac ATDPCs secreted proangiogenic factors under in vitro hypoxic conditions, suggesting a paracrine effect to promote local vascularization. Our results indicate that the population of progenitor cells isolated from human cardiac adipose tissue (cardiac ATDPCs) may be valid candidates for future use in cell therapy to regenerate injured myocardium. PMID:20713059

  10. Activated Factor X Induces Endothelial Cell Senescence Through IGFBP-5

    PubMed Central

    Sanada, Fumihiro; Taniyama, Yoshiaki; Muratsu, Jun; Otsu, Rei; Iwabayashi, Masaaki; Carracedo, Miguel; Rakugi, Hiromi; Morishita, Ryuichi

    2016-01-01

    Uncontrolled coagulation contributes to the pathophysiology of several chronic inflammatory diseases. In these conditions, senescent cells are often observed and is involved in the generation of inflammation. The coincidence of hyper-coagulation, cell senescence, and inflammation suggests the existence of a common underlying mechanism. Recent evidence indicates that activated coagulation factor X (FXa) plays a role in the processes beyond blood coagulation. This non-hematologic function entails the mediation of inflammation and tissue remodeling. We therefore tested the hypothesis that FXa induces cell senescence resulting in tissue inflammation and impaired tissue regeneration. Human umbilical vein endothelial cells were stimulated with FXa for 14 days. The proliferation of cells treated with FXa was significantly smaller, and the fraction of senescence-associated β-galactosidase-positive cells was increased as compared to the control group. RT-qPCR array revealed that FXa increased the expression of IGFBP-5, EGR-1, p53, and p16INK4a. Inhibition of FXa by a direct FXa inhibitor, rivaroxaban, or IGFBP-5 by siRNA decreased FXa-induced cell senescence, restoring cell proliferation. Moreover, in an ischemic hind limb mouse model, FXa inhibited neovascularization by endothelial progenitor cell. However, rivaroxaban significantly restored FXa-induced impaired angiogenesis. In summary, FXa induced endothelial cell senescence through IGFBP-5, resulting in impaired angiogenesis. PMID:27752126

  11. Design of biomimetic vascular grafts with magnetic endothelial patterning.

    PubMed

    Fayol, Delphine; Le Visage, Catherine; Ino, Julia; Gazeau, Florence; Letourneur, Didier; Wilhelm, Claire

    2013-01-01

    The development of small diameter vascular grafts with a controlled pluricellular organization is still needed for effective vascular tissue engineering. Here, we describe a technological approach combining a tubular scaffold and magnetically labeled cells to create a pluricellular and organized vascular graft, the endothelialization of which could be monitored by MRI prior to transplantation. A novel type of scaffold was developed with a tubular geometry and a porous bulk structure enabling the seeding of cells in the scaffold pores. A homogeneous distribution of human mesenchymal stem cells in the macroporous structure was obtained by seeding the freeze-dried scaffold with the cell suspension. The efficient covering of the luminal surface of the tube was then made possible thanks to the implementation of a magnetic-based patterning technique. Human endothelial cells or endothelial progenitors were magnetically labeled with iron oxide nanoparticles and successfully attracted to the 2-mm lumen where they attached and formed a continuous endothelium. The combination of imaging modalities [fluorescence imaging, histology, and 3D magnetic resonance imaging (MRI)] evidenced the integrity of the vascular construct. In particular, the observation of different cell organizations in a vascular scaffold within the range of resolution of single cells by 4.7 T MRI is reported. PMID:23295155

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

    PubMed Central

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

    2016-01-01

    The endothelial to haematopoietic transition (EHT) is a key developmental process where a drastic change of endothelial cell morphology leads to the formation of blood stem and progenitor cells during embryogenesis. As TGFβ signalling triggers a similar event during embryonic development called epithelial to mesenchymal transition (EMT), we hypothesised that TGFβ activity could play a similar role in EHT as well. We used the mouse embryonic stem cell differentiation system for in vitro recapitulation of EHT and performed gain and loss of function analyses of the TGFβ pathway. Quantitative proteomics analysis showed that TGFβ treatment during EHT increased the secretion of several proteins linked to the vascular lineage. Live cell imaging showed that TGFβ blocked the formation of round blood cells. Using gene expression profiling we demonstrated that the TGFβ signalling activation decreased haematopoietic genes expression and increased the transcription of endothelial and extracellular matrix genes as well as EMT markers. Finally we found that the expression of the transcription factor Sox17 was up-regulated upon TGFβ signalling activation and showed that its overexpression was enough to block blood cell formation. In conclusion we showed that triggering the TGFβ pathway does not enhance EHT as we hypothesised but instead impairs it. PMID:26891705

  13. Brain microglia and blood-derived macrophages: molecular profiles and functional roles in multiple sclerosis and animal models of autoimmune demyelinating disease.

    PubMed

    Raivich, Gennadij; Banati, Richard

    2004-11-01

    Microglia and macrophages, one a brain-resident, the other a mostly hematogenous cell type, represent two related cell types involved in the brain pathology in multiple sclerosis and its autoimmune animal model, the experimental allergic encephalomyelitis. Together, they perform a variety of different functions: they are the primary sensors of brain pathology, they are rapidly recruited to sites of infection, trauma or autoimmune inflammation in experimental allergic encephalomyelitis and multiple sclerosis and they are competent presenters of antigen and interact with T cells recruited to the inflamed CNS. They also synthesise a variety of molecules, such as cytokines (TNF, interleukins), chemokines, accessory molecules (B7, CD40), complement, cell adhesion glycoproteins (integrins, selectins), reactive oxygen radicals and neurotrophins, that could exert a damaging or a protective effect on adjacent axons, myelin and oligodendrocytes. The current review will give a detailed summary on their cellular response, describe the different classes of molecules expressed and their attribution to the blood derived or brain-resident macrophages and then discuss how these molecules contribute to the neuropathology. Recent advances using chimaeric and genetically modified mice have been particularly telling about the specific, overlapping and nonoverlapping roles of macrophages and microglia in the demyelinating disease. Interestingly, they point to a crucial role of hematogenous macrophages in initiating inflammation and myelin removal, and that of microglia in checking excessive response and in the induction and maintenance of remission.

  14. Pleiotrophin is involved in the amniotic epithelial cell-induced differentiation of human umbilical cord blood-derived mesenchymal stem cells into dopaminergic neuron-like cells.

    PubMed

    Yang, Shu; Xue, Dan-Dan; Wu, Bo; Sun, Hai-Mei; Li, Xiao-Shuang; Dong, Fang; Li, Wen-Shuai; Ji, Feng-Qing; Zhou, De-Shan

    2013-02-28

    We have reported that human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) are capable of differentiating into dopaminergic (DA) neuron-like cells upon being induced by amniotic epithelial cells (AECs). However, what factor(s) is involved in the differentiation process has not been explored out thoroughly. Because pleiotrophin (PTN) is known to exert important trophic effects on DA neurons, in the present study, we investigated whether PTN is released by AECs and whether it is involved in the differentiation of hUCB-MSCs into DA neuron-like cells. The expression and secretion of PTN by AECs were detected by immunofluorescence, RT-PCR and ELISA. The hUCB-MSCs were isolated and treated with AEC-conditioned medium (ACM) or recombinant human PTN. Compared to the controls, a higher proportion of treated cells differentiated into DA neuron-like cells, indicated by the increased expression of TH and DAT and the increased dopamine content. These results indicate that PTN released by AECs acts as a synergetic factor with other neurotrophic factors and is involved in the differentiation of hUCB-MSCs into DA neuron-like cells. We suggest that ACM, which contains PTN and other neurotrophic factors, could potentially be used as an agent to promote the differentiation of DA neuron-like cells from hUCB-MSCs for cell therapy of Parkinson's disease without creating legal or ethical issues.

  15. Mesenchymal Stromal/Stem Cell and Minocycline-Loaded Hydrogels Inhibit the Growth of Staphylococcus aureus that Evades Immunomodulation of Blood-Derived Leukocytes.

    PubMed

    Guerra, Alberto Daniel; Cantu, David Antonio; Vecchi, Joseph T; Rose, Warren E; Hematti, Peiman; Kao, Weiyuan John

    2015-05-01

    Mesenchymal stromal/stem cells (MSCs) have demonstrated favorable wound healing properties in addition to their differentiation capacity. MSCs encapsulated in biomaterials such as gelatin and polyethylene glycol (PEG) composite hydrogels have displayed an immunophenotype change that leads to the release of cytokines and growth factors to accelerate wound healing. However, therapeutic potential of implanted MSC-loaded hydrogels may be limited by non-specific protein adsorption that facilitates adhesion of bacterial pathogens such as planktonic Staphylococcus aureus (SA) to the surface with subsequent biofilm formation resistant to immune cell recognition and antibiotic activity. In this study, we demonstrate that blood-derived primary leukocytes and bone marrow-derived MSCs cannot inhibit colony-forming abilities of planktonic or biofilm-associated SA. However, we show that hydrogels loaded with MSCs and minocycline significantly inhibit colony-forming abilities of planktonic SA while maintaining MSC viability and multipotency. Our results suggest that minocycline and MSC-loaded hydrogels may decrease the bioburden of SA at implant sites in wounds, and may improve the wound healing capabilities of MSC-loaded hydrogels.

  16. Generation of a cord blood-derived Wilms Tumor 1 dendritic cell vaccine for AML patients treated with allogeneic cord blood transplantation

    PubMed Central

    de Haar, Colin; Plantinga, Maud; Blokland, Nina JG; van Til, Niek P; Flinsenberg, Thijs WH; Van Tendeloo, Viggo F; Smits, Evelien L; Boon, Louis; Spel, Lotte; Boes, Marianne; Boelens, Jaap Jan; Nierkens, Stefan

    2015-01-01

    The poor survival rates of refractory/relapsed acute myeloid leukemia (AML) patients after haematopoietic cell transplantation (HCT) requires the development of additional immune therapeutic strategies. As the elicitation of tumor-antigen specific cytotoxic T lymphocytes (CTLs) is associated with reduced relapses and enhanced survival, enhanced priming of these CTLs using an anti-AML vaccine may result in long-term immunity against AML. Cord blood (CB), as allogeneic HCT source, may provide a unique setting for such post-HCT vaccination, considering its enhanced graft-versus-leukemia (GvL) effects and population of highly responsive naïve T cells. It is our goal to develop a powerful and safe immune therapeutic strategy composed of CB-HCT followed by vaccination with CB CD34+-derived dendritic cells (DCs) presenting the oncoprotein Wilms Tumor-1 (WT1), which is expressed in AML-blasts in the majority of patients. Here, we describe the optimization of a clinically applicable DC culture protocol. This two-step protocol consisting of an expansion phase followed by the differentiation toward DCs, enables us to generate sufficient cord blood-derived DCs (CBDCs) in the clinical setting. At the end of the culture, the CBDCs exhibit a mature surface phenotype, are able to migrate, express tumor antigen (WT1) after electroporation with mRNA encoding the full-length WT1 protein, and stimulate WT1-specific T cells. PMID:26451309

  17. Human umbilical cord blood-derived mesenchymal stem cells improve neuropathology and cognitive impairment in an Alzheimer's disease mouse model through modulation of neuroinflammation.

    PubMed

    Lee, Hyun Ju; Lee, Jong Kil; Lee, Hyun; Carter, Janet E; Chang, Jong Wook; Oh, Wonil; Yang, Yoon Sun; Suh, Jun-Gyo; Lee, Byoung-Hee; Jin, Hee Kyung; Bae, Jae-Sung

    2012-03-01

    Human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSC) have a potential therapeutic role in the treatment of neurological disorders, but their current clinical usage and mechanism of action has yet to be ascertained in Alzheimer's disease (AD). Here we report that hUCB-MSC transplantation into amyloid precursor protein (APP) and presenilin1 (PS1) double-transgenic mice significantly improved spatial learning and memory decline. Furthermore, amyloid-β peptide (Aβ) deposition, β-secretase 1 (BACE-1) levels, and tau hyperphosphorylation were dramatically reduced in hUCB-MSC transplanted APP/PS1 mice. Interestingly, these effects were associated with reversal of disease-associated microglial neuroinflammation, as evidenced by decreased microglia-induced proinflammatory cytokines, elevated alternatively activated microglia, and increased anti-inflammatory cytokines. These findings lead us to suggest that hUCB-MSC produced their sustained neuroprotective effect by inducing a feed-forward loop involving alternative activation of microglial neuroinflammation, thereby ameliorating disease pathophysiology and reversing the cognitive decline associated with Aβ deposition in AD mice.

  18. Autocrine Action of Thrombospondin-2 Determines the Chondrogenic Differentiation Potential and Suppresses Hypertrophic Maturation of Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells.

    PubMed

    Jeong, Sang Young; Ha, Jueun; Lee, Miyoung; Jin, Hye Jin; Kim, Dong Hyun; Choi, Soo Jin; Oh, Wonil; Yang, Yoon Sun; Kim, Jae-Sung; Kim, Byung-Gyu; Chang, Jeong Ho; Cho, Dong-Hyung; Jeon, Hong Bae

    2015-11-01

    Previous studies have shown that mesenchymal stem cell (MSC)-based therapies have varying efficacies for the treatment of various diseases, including cartilage defects. In this study, we demonstrated that the chondrogenic differentiation potential of human umbilical cord blood-derived MSCs (hUCB-MSCs) obtained from different individual donors varies, and we investigated the molecular basis for this variation. Microarray gene expression analysis identified thrombospondin-2 (TSP2) as a candidate gene underlying the interindividual variation in the chondrogenic differentiation potential of hUCB-MSCs. To assess the association between TSP-2 and the differentiation potential, we evaluated chondrogenic differentiation of hUCB-MSCs treated with TSP2 siRNA. In addition, we studied the effect of supplementing exogenous recombinant TSP-2 on TSP2 siRNA-treated hUCB-MSCs. We found that TSP-2 autocrinally promoted chondrogenic differentiation of hUCB-MSCs via the Notch signaling pathway, which was confirmed in MSCs from other sources such as bone marrow and adipose tissue. Interestingly, we observed that TSP-2 attenuated hypertrophy, which inevitably occurs during chondrogenic differentiation of hUCB-MSCs. Our findings indicated that the variable chondrogenic differentiation potential of MSCs obtained from different donors is influenced by the TSP-2 level in the differentiating cells. Thus, the TSP-2 level can be used as a marker to select MSCs with superior chondrogenic differentiation potential for use in cartilage regeneration therapy. PMID:26235673

  19. Human progenitor cell recruitment via SDF-1α coacervate-laden PGS vascular grafts.

    PubMed

    Lee, Kee-Won; Johnson, Noah R; Gao, Jin; Wang, Yadong

    2013-12-01

    Host cell recruitment is crucial for vascular graft remodeling and integration into the native blood vessel; it is especially important for cell-free strategies which rely on host remodeling. Controlled release of growth factors from vascular grafts may enhance host cell recruitment. Stromal cell-derived factor (SDF)-1α has been shown to induce host progenitor cell migration and recruitment; however, its potential in regenerative therapies is often limited due to its short half-life in vivo. This report describes a coacervate drug delivery system for enhancing progenitor cell recruitment into an elastomeric vascular graft by conferring protection of SDF-1α. Heparin and a synthetic polycation are used to form a coacervate, which is incorporated into poly(glycerol sebacate) (PGS) scaffolds. In addition to protecting SDF-1α, the coacervate facilitates uniform scaffold coating. Coacervate-laden scaffolds have high SDF-1α loading efficiency and provide sustained release under static and physiologically-relevant flow conditions with minimal initial burst release. In vitro assays showed that coacervate-laden scaffolds enhance migration and infiltration of human endothelial and mesenchymal progenitor cells by maintaining a stable SDF-1α gradient. These results suggest that SDF-1α coacervate-laden scaffolds show great promise for in situ vascular regeneration.

  20. PDGFRα demarcates the cardiogenic clonogenic Sca1+ stem/progenitor cell in adult murine myocardium.

    PubMed

    Noseda, Michela; Harada, Mutsuo; McSweeney, Sara; Leja, Thomas; Belian, Elisa; Stuckey, Daniel J; Abreu Paiva, Marta S; Habib, Josef; Macaulay, Iain; de Smith, Adam J; al-Beidh, Farah; Sampson, Robert; Lumbers, R Thomas; Rao, Pulivarthi; Harding, Sian E; Blakemore, Alexandra I F; Jacobsen, Sten Eirik; Barahona, Mauricio; Schneider, Michael D

    2015-05-18

    Cardiac progenitor/stem cells in adult hearts represent an attractive therapeutic target for heart regeneration, though (inter)-relationships among reported cells remain obscure. Using single-cell qRT-PCR and clonal analyses, here we define four subpopulations of cardiac progenitor/stem cells in adult mouse myocardium all sharing stem cell antigen-1 (Sca1), based on side population (SP) phenotype, PECAM-1 (CD31) and platelet-derived growth factor receptor-α (PDGFRα) expression. SP status predicts clonogenicity and cardiogenic gene expression (Gata4/6, Hand2 and Tbx5/20), properties segregating more specifically to PDGFRα(+) cells. Clonal progeny of single Sca1(+) SP cells show cardiomyocyte, endothelial and smooth muscle lineage potential after cardiac grafting, augmenting cardiac function although durable engraftment is rare. PDGFRα(-) cells are characterized by Kdr/Flk1, Cdh5, CD31 and lack of clonogenicity. PDGFRα(+)/CD31(-) cells derive from cells formerly expressing Mesp1, Nkx2-5, Isl1, Gata5 and Wt1, distinct from PDGFRα(-)/CD31(+) cells (Gata5 low; Flk1 and Tie2 high). Thus, PDGFRα demarcates the clonogenic cardiogenic Sca1(+) stem/progenitor cell.

  1. PDGFRα demarcates the cardiogenic clonogenic Sca1+ stem/progenitor cell in adult murine myocardium

    PubMed Central

    Noseda, Michela; Harada, Mutsuo; McSweeney, Sara; Leja, Thomas; Belian, Elisa; Stuckey, Daniel J.; Abreu Paiva, Marta S.; Habib, Josef; Macaulay, Iain; de Smith, Adam J.; al-Beidh, Farah; Sampson, Robert; Lumbers, R. Thomas; Rao, Pulivarthi; Harding, Sian E.; Blakemore, Alexandra I. F.; Eirik Jacobsen, Sten; Barahona, Mauricio; Schneider, Michael D.

    2015-01-01

    Cardiac progenitor/stem cells in adult hearts represent an attractive therapeutic target for heart regeneration, though (inter)-relationships among reported cells remain obscure. Using single-cell qRT–PCR and clonal analyses, here we define four subpopulations of cardiac progenitor/stem cells in adult mouse myocardium all sharing stem cell antigen-1 (Sca1), based on side population (SP) phenotype, PECAM-1 (CD31) and platelet-derived growth factor receptor-α (PDGFRα) expression. SP status predicts clonogenicity and cardiogenic gene expression (Gata4/6, Hand2 and Tbx5/20), properties segregating more specifically to PDGFRα+ cells. Clonal progeny of single Sca1+ SP cells show cardiomyocyte, endothelial and smooth muscle lineage potential after cardiac grafting, augmenting cardiac function although durable engraftment is rare. PDGFRα− cells are characterized by Kdr/Flk1, Cdh5, CD31 and lack of clonogenicity. PDGFRα+/CD31− cells derive from cells formerly expressing Mesp1, Nkx2-5, Isl1, Gata5 and Wt1, distinct from PDGFRα−/CD31+ cells (Gata5 low; Flk1 and Tie2 high). Thus, PDGFRα demarcates the clonogenic cardiogenic Sca1+ stem/progenitor cell. PMID:25980517

  2. Bone Marrow-Derived Progenitor Cells Are Functionally Impaired in Ischemic Heart Disease.

    PubMed

    Nollet, Evelien; Hoymans, Vicky Y; Rodrigus, Inez R; De Bock, Dina; Dom, Marc; Vanassche, Bruno; Van Hoof, Viviane O M; Cools, Nathalie; Van Ackeren, Katrijn; Wouters, Kristien; Vermeulen, Katrien; Vrints, Christiaan J; Van Craenenbroeck, Emeline M

    2016-08-01

    To determine whether the presence of ischemic heart disease (IHD) per se, or rather the co-presence of heart failure (HF), is the primum movens for less effective stem cell products in autologous stem cell therapy, we assessed numbers and function of bone marrow (BM)-derived progenitor cells in patients with coronary artery disease (n = 17), HF due to ischemic cardiomyopathy (n = 8), non-ischemic HF (n = 7), and control subjects (n = 11). Myeloid and erythroid differentiation capacity of BM-derived mononuclear cells was impaired in patients with underlying IHD but not with non-ischemic HF. Migration capacity decreased with increasing IHD severity. Hence, IHD, with or without associated cardiomyopathy, is an important determinant of progenitor cell function. No depletion of hematopoietic and endothelial progenitor cells (EPC) within the BM was observed, while circulating EPC numbers were increased in the presence of IHD, suggesting active recruitment. The observed myelosuppression was not driven by inflammation and thus