Germana Castelli

Istituto Superiore di Sanità, Roma, Latium, Italy

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Publications (14)75.43 Total impact

  • Elvira Pelosi, Germana Castelli, Ugo Testa
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    ABSTRACT: The studies carried out during the last two decades have represented a great effort in trying to identify and define cell populations endowed with the phenotypic and functional properties of endothelial progenitors. From these studies a scenario now emerges indicating that in the blood there are very rare endothelial progenitor cells, called endothelial colony-forming cells (ECFCs) or late outgrowth endothelial cells, not originated from bone marrow, capable of generating phenotypically and functionally competent endothelial cells, capable to be incorporated in vivo into growing vessels. ECFCs are present in the circulation as well as cells resident in the vascular endothelial intima. In addition to these progenitors, there are some hematopoietic progenitor cells capable of generating a monocytic cell progeny exerting a pro-angiogenic activity in vivo, but unable to be directly incorporated into growing vessels. These cells exert a pro-angiogenic effect in vivo through a paracrine mechanism based on the secretion of growth factors and cytokines.
    Blood Cells Molecules and Diseases 12/2013; · 2.26 Impact Factor
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    ABSTRACT: : HIV infection affects dendritic cells (DCs) number, maturation, and function although the cause remains largely unknown. Purified CD34 hematopoietic progenitor cells (HPCs) obtained from bone marrow of chronic HIV-infected patients were investigated for the differentiative capability toward mature DCs. HIV, although not in active replication, was found able to impair CD34 HPC differentiation into mature DCs. These results suggest that DCs impairment found in HIV-infected patients may be related to a failure by bone marrow CD34 HPCs to produce an adequate number of DCs.
    JAIDS Journal of Acquired Immune Deficiency Syndromes 12/2013; 64(4):342-4. · 4.65 Impact Factor
  • Elvira Pelosi, Germana Castelli, Ugo Testa
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    ABSTRACT: Cord blood (CB) is a rich source of hematopoietic stem cells (HSCs) and for this reason CB transplantation has been used successfully for the treatment of some malignant and nonmalignant diseases. However, this technique is limited by the relatively low number of HSCs present in each CB unit and by the delayed engraftment of platelets and neutrophils. To bypass these obstacles efforts have been made to develop strategies to expand CB HSCs in vitro for transplantation. CB is also an important source of other stem cells, including endothelial progenitors, mesenchymal stem cells (MSCs), very small embryonic/epiblast-like (VSEL) stem cells, and unrestricted somatic stem cells (USSC), potentially suitable for use in regenerative medicine. For some of these stem cell populations, such as MSCs, clinical studies have been started and for other stem cell populations potential clinical applications have been identified and clinical studies will follow. In addition to CB, other parts of umbilical cord, such as the Wharton's jelly, or tissues strictly linked such as the placenta are also rich sources of stem cells.
    Blood Cells Molecules and Diseases 03/2012; 49(1):20-8. · 2.26 Impact Factor
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    ABSTRACT: The tyrosine kinase Tie-2 and its ligands Angiopoietins (Angs) transduce critical signals for angiogenesis in endothelial cells. This receptor and Ang-1 are coexpressed in hematopoietic stem cells and in a subset of megakaryocytes, though a possible role of angiopoietins in megakaryocytic differentiation/proliferation remains to be demonstrated. To investigate a possible effect of Ang-1/Ang-2 on megakaryocytic proliferation/differentiation we have used both normal CD34(+) cells induced to megakaryocytic differentiation and the UT7 cells engineered to express the thrombopoietin receptor (TPOR, also known as c-mpl, UT7/mpl). Our results indicate that Ang-1/Ang-2 may have a role in megakaryopoiesis. Particularly, Ang-2 is predominantly produced and released by immature normal megakaryocytic cells and by undifferentiated UT7/mpl cells and slightly stimulated TPO-induced cell proliferation. Ang-1 production is markedly induced during megakaryocytic differentiation/maturation and potentiated TPO-driven megakaryocytic differentiation. Blocking endogenously released angiopoietins partially inhibited megakaryocytic differentiation, particularly for that concerns the process of polyploidization. According to these data it is suggested that an autocrine angiopoietin/Tie-2 loop controls megakaryocytic proliferation and differentiation.
    PLoS ONE 01/2012; 7(7):e39796. · 3.53 Impact Factor
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    ABSTRACT: Embryologic and genetic evidence suggest a common origin of haematopoietic and endothelial lineages. In the murine embryo, recent studies indicate the presence of haemogenic endothelium and of a common haemato-endothelial precursor, the haemangioblast. Conversely, so far, little evidence supports the presence of haemogenic endothelium and haemangioblasts in later stages of development. Our studies indicate that human cord blood haematopoietic progenitors (CD34+45+144-), triggered by murine hepatocyte conditioned medium, differentiate into adherent proliferating endothelial precursors (CD144+CD105+CD146+CD31+CD45-) capable of functioning as haemogenic endothelium. These cells, proven to give rise to functional vasculature in vivo, if further instructed by haematopoietic growth factors, first switch to transitional CD144+45+ cells and then to haematopoietic cells. These results highlight the plasticity of haemato-endhothelial precursors in human post-natal life. Furthermore, these studies may provide highly enriched populations of human post-fetal haemogenic endothelium, paving the way for innovative projects at a basic and possibly clinical level.
    PLoS ONE 01/2012; 7(12):e51109. · 3.53 Impact Factor
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    ABSTRACT: FMS-related tyrosine kinase 3 (FLT3) mutations are found in 30% of cases of acute myeloid leukaemia (AML). In addition, recent studies have lead to the identification of about 10-15% of AML patients displaying high expression of FLT3, not associated with mutations of the receptor (FLT3 Wild-type High, FLT3WTH). These AMLs, as well as those displaying internal tandem duplication (ITD) are associated with an unfavourable prognosis. However, the biological features of these AMLs are poorly characterized. The present study explored the immunophenotypic features of FLT3WTH AMLs in 94 de novo cases of AML. The levels of FLT3 expression, as assessed by flow cytometry and FLT3 mutational status, was used to identify four AML subgroups: FLT3WTH (14/94); FLT3 Wild-type low (FLT3WTL, 48/94); FLT3 internal tandem duplication (FLT3ITD 26/94); FLT3 aspartic acid 835 (FLT3D835, 6/94). FLT3WTH and FLT3ITD were characterized by: high white blast cell counts; predominance of M4 and M5 French-American-British classification subtypes and associated expression of myelo-monocytic markers; high expression of CD123 and TRAIL-Rs; high expression of receptors for angiogenic growth factors. Addition of FLT3 Ligand to human CD34(+) or monocytic cells stimulated CD123 and TRAIL-R expression. These findings are of potential value for the development of new therapeutic strategies.
    British Journal of Haematology 02/2011; 153(1):33-42. · 4.94 Impact Factor
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    ABSTRACT: The human hemoglobin switch (HbF-->HbA) takes place in the peri/post-natal period. In adult life, however, the residual HbF (<1%) may be partially reactivated by chemical inducers and/or cytokines such as the kit ligand (KL). MicroRNAs (miRs) play a pivotal role in normal hematopoiesis: downmodulation of miR-221/222 stimulates human erythropoietic proliferation through upmodulation of the kit receptor. We have explored the possible role of kit/KL in perinatal Hb switching by evaluating: i) the expression levels of both kit and kit ligand on CD34(+) cells and in plasma isolated from pre-, mid- and full-term cord blood samples; ii) the reactivation of HbF synthesis in KL-treated unilineage erythroid cell cultures; iii) the functional role of miR-221/222 in HbF production. In perinatal life, kit expression showed a gradual decline directly correlated to the decrease of HbF (from 80-90% to <30%). Moreover, in full-term cord blood erythroid cultures, kit ligand induced a marked increase of HbF (up to 80%) specifically abrogated by addition of the kit inhibitor imatinib, thus reversing the Hb switch. MiR-221/222 expression exhibited rising levels during peri/post-natal development. In functional studies, overexpression of these miRs in cord blood progenitors caused a remarkable decrease in kit expression, erythroblast proliferation and HbF content, whereas their suppression induced opposite effects. Our studies indicate that human perinatal Hb switching is under control of the kit receptor/miR 221-222 complex. We do not exclude, however, that other mechanisms (i.e. glucocorticoids and the HbF inhibitor BCL11A) may also contribute to the peri/post-natal Hb switch.
    Haematologica 03/2010; 95(8):1253-60. · 5.94 Impact Factor
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    ABSTRACT: It is generally conceded that selective combinations of transcription factors determine hematopoietic lineage commitment and differentiation. Here we show that in normal human hematopoiesis the transcription factor nuclear factor I-A (NFI-A) exhibits a marked lineage-specific expression pattern: it is upmodulated in the erythroid (E) lineage while fully suppressed in the granulopoietic (G) series. In unilineage E culture of hematopoietic progenitor cells (HPCs), NFI-A overexpression or knockdown accelerates or blocks erythropoiesis, respectively: notably, NFI-A overexpression restores E differentiation in the presence of low or minimal erythropoietin stimulus. Conversely, NFI-A ectopic expression in unilineage G culture induces a sharp inhibition of granulopoiesis. Finally, in bilineage E + G culture, NFI-A overexpression or suppression drives HPCs into the E or G differentiation pathways, respectively. These NFI-A actions are mediated, at least in part, by a dual and opposite transcriptional action: direct binding and activation or repression of the promoters of the beta-globin and G-CSF receptor gene, respectively. Altogether, these results indicate that, in early hematopoiesis, the NFI-A expression level acts as a novel factor channeling HPCs into either the E or G lineage.
    Blood 07/2009; 114(9):1753-63. · 9.78 Impact Factor
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    ABSTRACT: The promyelocytic leukemia zinc-finger protein (PLZF) is a transcription factor and c-kit is a receptor tyrosine kinase associated with human disease, particularly in hematopoietic cells. MicroRNAs (miRs) are post-transcriptional regulators of gene expression, and c-kit has been described as a target of miRs-221 and -222 in erythropoiesis. In the present study, we identified c-kit as a target of PLZF in normal and leukemic cells. Particularly, in erythropoietic (E) culture of CD34(+) progenitors, PLZF is downregulated, whereas c-kit expression at both the mRNA and protein levels inversely increases during the first days of E differentiation. In functional experiments, PLZF transfection induces c-kit downregulation, inhibits E proliferation and delays differentiation, whereas PLZF knockdown induces opposite effects, independently of miRs-221 and -222 expression. The inverse correlation between PLZF and c-kit expression was found in normal CD34(+)38(+/-) hematopoietic progenitor/stem cells and in acute myeloid leukemias of M0/M1 French-American-British subtypes, suggesting that the control of PLZF on c-kit expression may be crucial at the level of the stem cell/progenitor compartment. Altogether, our data indicate a new mechanism of regulation of c-kit expression that involves a transcriptional control by PLZF in CD34(+) cells and early erythropoiesis.
    Oncogene 06/2009; 28(23):2276-88. · 8.56 Impact Factor
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    ABSTRACT: MicroRNAs are small non-coding RNAs that regulate gene expression through mRNA degradation or translational inhibition. MicroRNAs are emerging as key regulators of normal hematopoiesis and hematologic malignancies. Several miRNAs are differentially expressed during hematopoiesis and their specific expression regulates key functional proteins involved in hematopoietic lineage differentiation. This study focused on the functional role of microRNA-223 (miR-223) on erythroid differentiation. Purified cord blood CD34+ hematopoietic progenitor cells were grown in strictly controlled conditions in the presence of saturating dosage of erythropoietin to selectively induce erythroid differentiation. The effects of enforced expression of miR-223 in unilin-eage erythroid cultures were evaluated in liquid phase culture experiments and clonogenic studies. In unilineage erythroid culture of cord blood CD34+ hematopoietic progenitor cells miR-223 is down-regulated, whereas LMO2, an essential protein for erythroid differentiation, is up-regulated. Functional studies showed that enforced expression of miR-223 reduces the mRNA and protein levels of LMO2, by binding to LMO2 3' UTR, and impairs differentiation of erythroid cells. Accordingly, knockdown of LMO2 by short interfering RNA mimics the action of miR-223. Furthermore, hematopoietic progenitor cells transduced with miR-223 showed a significant reduction of their erythroid clonogenic capacity, suggesting that downmodulation of this miRNA is required for erythroid progenitor recruitment and commitment. These results show that the decline of miR-223 is an important event for erythroid differentiation that leads to the expansion of erythroblast cells at least partially mediated by unblocking LMO2 protein expression.
    Haematologica 05/2009; 94(4):479-86. · 5.94 Impact Factor
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    ABSTRACT: Mesenchymal stromal cells (MSCs) represent a bone marrow (BM) population, classically defined by five functional properties: extensive proliferation, ability to differentiate into osteoblasts, chondrocytes, adipocytes, and stromal cells-supporting hematopoiesis. However, research progress in this area has been hampered by lack of suitable markers and standardized procedures for MSC isolation. We have isolated a CD146(+) multipotent MSC population from 20 human BM donors displaying the phenotype of self-renewing osteoprogenitors; an extensive 12-week proliferation; and the ability to differentiate in osteoblasts, chondrocytes, adipocytes, and stromal cells supporting hematopoiesis. Furthermore, the CD146(+) MSCs secrete a complex combination of growth factors (GFs) controlling hematopoietic stem cells (HSCs) function, while providing a >2-log increase in the long-term culture (LTC) colony output in 8-week LTC over conventional assays. The hematopoietic stromal function exhibited by the MSCs was further characterized by manipulating LTCs with the chemical inhibitors Imatinib or SU-5416, targeting two GF receptors (GFRs), KIT or VEGFR2/1, respectively. Both treatments similarly impaired LTC colony output, indicating key roles for these two GF/GFR interactions to support LTC-initiating cell activity. CD146(+) MSCs may thus represent a tool to explore the MSC-HSC cross-talk in an in vitro surrogate model for HSC "niches," and for regenerative therapy studies. In addition, the MSC microRNA (miRNA) expression profile was analyzed by microarrays in both basic conditions and chondrogenic differentiation. Our analysis revealed that several miRNAs are modulated during chondrogenesis, and many of their putative targets are genes involved in chondrogenic differentiation.
    Experimental Hematology 08/2008; 36(8):1035-46. · 2.91 Impact Factor
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    ABSTRACT: We have developed a new culture system whereby human hematopoietic progenitors purified from adult peripheral blood extensively proliferate and gradually differentiate into >95% pure monocytic (Mo) cells. At all developmental stages treatment with interleukin (IL)-4+granulocyte-macrophage colony-stimulating factor or IL-4+c-Kit-ligand+FLT-3 ligand switched the Mo precursors into dendritic cells (DCs). The switching capacity decreased only at the end of the culture, when most Mo cells matured to macrophages. Moreover, the Mo precursors were highly susceptible to transduction with lentiviral vectors: once switched to DCs, they maintained the transgene expression, as well as the phenotype and function of the DC lineage. Our results provide new insight into the potential role of the Mo lineage as a reservoir of DCs in vivo. Furthermore, the methodology for transduction of Mo precursors provides a tool to generate genetically modified, normally functioning DCs potentially useful for immunotherapy.
    Cell Death and Differentiation 02/2006; 13(2):250-9. · 8.37 Impact Factor
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    ABSTRACT: The expression/function of vascular endothelial growth factor (VEGF) receptors (VEGFR1/Flt1 and VEGFR2/KDR/Flk1) in hematopoiesis is under scrutiny. We have investigated the expression of Flt1 and kinase domain receptor (KDR) on hematopoietic precursors, as evaluated in liquid culture of CD34(+) hematopoietic progenitor cells (HPCs) induced to unilineage differentiation/maturation through the erythroid (E), megakaryocytic (Mk), granulocytic (G), or monocytic (Mo) lineage. KDR, expressed on 0.5% to 1.5% CD34(+) cells, is rapidly downmodulated on induction of differentiation. Similarly, Flt1 is present at very low levels in HPCs and is downmodulated in E and G lineages; however, Flt1 is induced in the precursors of both Mo and Mk series; ie, its level progressively increases during Mo maturation, and it peaks at the initial-intermediate culture stages in the Mk lineage. Functional experiments indicate that Mk and E, but not G and Mo, precursors release significant amounts of VEGF in the culture medium, particularly at low O(2) levels. The functional role of VEGF release on Mk maturation is indicated by 2 series of observations. (1) Molecules preventing the VEGF-Flt1 interaction on the precursor membrane (eg, soluble Flt1 receptors) significantly inhibit Mk polyploidization. (2) Addition of exogenous VEGF or placenta growth factor (PlGF) markedly potentiates Mk maturation. Conversely, VEGF does not modify Mo differentiation/maturation. Altogether, our results suggest that in the hematopoietic microenvironment an autocrine VEGF loop contributes to optimal Mk maturation through Flt1. A paracrine loop involving VEGF release by E precursors may also operate. Similarly, recent studies indicate that an autocrine loop involving VEGF and Flt1/Flk1 receptors mediates hematopoietic stem cell survival and differentiation.
    Blood 03/2003; 101(4):1316-23. · 9.78 Impact Factor
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    ABSTRACT: Dendritic cells (DCs) perform a basic role in the immune system by allowing the initiation of the primary T-cell-dependent immune response. Given previous indirect evidence that DC maturation and function are impaired by HIV, we have developed an in vitro culture system in order to verify the effect of HIV infection on DC function during the development from hematopoietic progenitors. Considering that monocytic (Mo) differentiating cells efficiently replicate monocytotropic HIV, we examined whether HIV-infected monocytic precursors (MoP) were able to generate functional DCs. CD34+ hematopoietic progenitor cells (HPCs) were induced along Mo differentiative pathway in liquid cultures and at an early stage of culture, MoP were infected with M-tropic BaL HIV strain, and after 2 days they were switched to DC differentiation with GM-CSF and IL-4. Derived DCs were actively infected, as detected by HIV-p24 production. HIV did not significantly affect cell viability, but induced a reduction in cell proliferation and an inefficient functional activity in terms of uptake capability and stimulation of allogenic T cells. These results indicate that HIV-infected MoP lost the capacity to generate functional DCs, and this may represent one of the many mechanisms of immunosuppression exploited by HIV.
    International journal of immunopathology and pharmacology 26(3):717-24. · 2.99 Impact Factor

Publication Stats

271 Citations
75.43 Total Impact Points

Institutions

  • 2003–2013
    • Istituto Superiore di Sanità
      • Department of Haematology, Oncology and Molecular Medicine
      Roma, Latium, Italy
  • 2009
    • Sapienza University of Rome
      • Unit of Histology and Medical Embryology
      Roma, Latium, Italy