A Balduini

Publications (10)47.43 Total impact

• Article: Ubiquitin/proteasome-rich particulate cytoplasmic structures (PaCSs) in the platelets and megakaryocytes of ANKRD26-related thrombo-cytopenia.

  V Necchi, A Balduini, P Noris, S Barozzi, P Sommi, C di Buduo, C L Balduini, E Solcia, A Pecci
  [show abstract] [hide abstract]
  ABSTRACT: ANKRD26-related thrombocytopenia (ANKRD26-RT) is an autosomal-dominant thrombocytopenia caused by mutations in the 5'UTR of the ANKRD26 gene. ANKRD26-RT is characterised by dysmegakaryopoiesis and an increased risk of leukaemia. PaCSs are novel particulate cytoplasmic structures with selective immunoreactivity for polyubiquitinated proteins and proteasome that have been detected in a number of solid cancers, in the epithelia of Helicobacter pylori gastritis and related preneoplastic lesions, and in the neutrophils of Schwachman-Diamond syndrome, a genetic disease with neutropenia and increased leukaemia risk. We searched for PaCSs in blood cells from 14 consecutive patients with ANKRD26-RT. Electron microscopy combined with immunogold staining for polyubiquitinated proteins, 20S and 19S proteasome showed PaCSs in most ANKRD26-RT platelets, as in a restricted minority of platelets from healthy controls and from subjects with other inherited or immune thrombocytopenias. In ANKRD26-RT platelets, the PaCS amount exceeded that of control platelets by a factor of 5 (p<0.0001). Immunoblotting showed that the higher PaCS number was associated with increased amounts of polyubiquitinated proteins and proteasome in ANKRD26-RT platelets. PaCSs were also extensively represented in ANKRD26-RT megakaryocytes, but not in healthy control megakaryocytes, and were absent in other ANKRD26-RT and control blood cells. Therefore, large amounts of PaCSs are a characteristic feature of ANKRD26-RT platelets and megakaryocytes, although these novel cell components are also present in a small subpopulation of normal platelets. The widespread presence of PaCSs in inherited diseases with increased leukaemia risk, as well as in solid neoplasms and their preneoplastic lesions, suggests a link of these structures with oncogenesis.
  Thrombosis and Haemostasis 12/2012; 109(2). · 5.04 Impact Factor
• Article: Proplatelet formation in heterozygous Bernard-Soulier syndrome type Bolzano.

  A Balduini, A Malara, A Pecci, S Badalucco, V Bozzi, I Pallotta, P Noris, M Torti, C L Balduini
  [show abstract] [hide abstract]
  ABSTRACT: Although mutations of GPIb alpha are among the most frequent causes of inherited platelet disorders, the mechanisms for the onset of thrombocytopenia and platelet macrocytosis are still poorly defined. In this work we analyzed in vitro megakaryocyte differentiation and proplatelet formation in six subjects heterozygous for the Ala156Val mutation in the GPIb alpha (Bolzano mutation). Human megakaryocytes were obtained by differentiation of patient cord blood-derived CD34(+) cells and peripheral blood-derived CD45(+) cells. Proplatelet formation was evaluated by phase contrast and fluorescence microscopy. Megakaryocyte differentiation from both cord blood (one patient) and peripheral blood (five patients) was comparable to controls. However, proplatelet formation was reduced by about 50% with respect to controls. An identical defect of proplatelet formation was observed when megakaryocytes were plated on fibrinogen, von Willebrand factor or grown in suspension. Morphological evaluation of proplatelet formation revealed an increased size of proplatelet tips, which was consistent with the increased diameters of patients' blood platelets. Moreover, alpha-tubulin distribution within proplatelets was severely deranged. Megakaryocytes from patients carrying a Bolzano allele of GPIb alpha display both quantitative and qualitative abnormalities of proplatelet formation in vitro. These results suggest that a defect of platelet formation contributes to macrothrombocytopenia associated to the Bolzano mutation, and indicate a key role for GPIb alpha in proplatelet formation.
  Journal of Thrombosis and Haemostasis 01/2009; 7(3):478-84. · 5.73 Impact Factor
• Article: Adhesive receptors, extracellular proteins and myosin IIA orchestrate proplatelet formation by human megakaryocytes.

  A Balduini, I Pallotta, A Malara, P Lova, A Pecci, G Viarengo, C L Balduini, M Torti
  [show abstract] [hide abstract]
  ABSTRACT: Megakaryocytes release platelets from the tips of cytoplasmic extensions, called proplatelets. In humans, the regulation of this process is still poorly characterized. To analyse the regulation of proplatelet formation by megakaryocyte adhesion to extracellular adhesive proteins through different membrane receptors. Human megakaryocytes were obtained by differentiation of cord blood-derived CD34(+) cells, and proplatelet formation was evaluated by phase contrast and fluorescence microscopy. We found that human megakaryocytes extended proplatelets in a time-dependent manner. Adhesion to fibrinogen, fibronectin or von Willebrand factor (VWF) anticipated the development of proplatelets, but dramatically limited both amplitude and duration of the process. Type I, but not type III or type IV, collagen totally suppressed proplatelet extension, and this effect was overcome by the myosin IIA antagonist blebbistatin. Integrin alphaIIbbeta3 was essential for megakaryocyte spreading on fibrinogen or VWF, but was not required for proplatelet formation. In contrast, proplatelet formation was prevented by blockade of GPIb-IX-V, or upon cleavage of GPIbalpha by the metalloproteinase mocarhagin. Membrane-associated VWF was detected exclusively on proplatelet-forming megakaryocytes, but not on round mature cells that do not extend proplatelets. Our findings show that proplatelet formation in human megakaryocytes undergoes a complex spatio-temporal regulation orchestrated by adhesive proteins, GPIb-IX-V and myosin IIA.
  Journal of Thrombosis and Haemostasis 09/2008; 6(11):1900-7. · 5.73 Impact Factor
• Article: Cord blood in vitro expanded CD41 cells: identification of novel components of megakaryocytopoiesis.

  A Balduini, M d'Apolito, D Arcelli, V Conti, A Pecci, D Pietra, M Danova, F Benvenuto, C Perotti, L Zelante, S Volinia, C L Balduini, A Savoia
  [show abstract] [hide abstract]
  ABSTRACT: Megakaryopoiesis represents a multi-step, often unclear, process leading to commitment, differentiation, and maturation of megakaryocytes (MKs) that release platelets. To identify the novel genes that might help to clarify the molecular mechanisms of megakaryocytopoiesis and be regarded as potential candidates of inherited platelet defects, global gene expression of hematopoietic lineages was carried out. Human cord blood was used to purify CD34+ stem cells and in vitro expand CD41+ cells and burst-forming unit-erythroid (BFU-E). We investigated the expression profiles of these three hematopoietic lineages in the Affymetrix system and selected genes specifically expressed in MKs by comparing transcripts of the different lineages using the dchip and pam algorithms. A detailed characterization of MK population showed that 99% of cells expressed the CD41 antigen whereas 73% were recognizable as terminally differentiated fetal MKs. The profile of these cells was compared with that of CD34+ cells and BFU-E allowing us to select 70 transcripts (MK-core), which represent not only the genes with a well-known function in MKs, but also novel genes never detected or characterized in these cells. Moreover, the specific expression was confirmed at both RNA and protein levels, thus validating the 'MK-core' isolated by informatics tools. This is a global gene expression that for the first time depicts a well-characterized population of cord blood-derived fetal MKs. Novel genes have been detected, such as those encoding components of the extracellular matrix and basal membrane, which have been found in the cytoplasm of Mks, suggesting that new physiological aspects of MKs should be studied.
  Journal of Thrombosis and Haemostasis 05/2006; 4(4):848-60. · 5.73 Impact Factor
• Article: Altered cytoskeleton organization in platelets from patients with MYH9-related disease.

  I Canobbio, P Noris, A Pecci, A Balduini, C L Balduini, M Torti
  [show abstract] [hide abstract]
  ABSTRACT: MYH9-related disease (MYH9-RD) is an autosomal dominant disorder deriving from mutations in the MYH9 gene encoding for the heavy chain of non-muscle myosin IIA, and characterized by thrombocytopenia and giant platelets. Isoform IIA of myosin is the only one expressed in platelets, but the possibility that MYH9 mutations affect the organization of contractile structures in these blood elements has never been investigated. In this work we have analyzed the composition and the agonist-induced reorganization of the platelet cytoskeleton from seven MYH9-RD patients belonging to four different families. We found that an increased amount of myosin was constitutively associated with actin in the cytoskeleton of resting MYH9-RD platelets. Upon platelet stimulation, an impaired increase in the total cytoskeletal proteins was observed. Moreover, selected membrane glycoproteins, tyrosine kinases, and small GTPases failed to interact with the cytoskeleton in agonist-stimulated MYH9-RD platelets. These results demonstrate for the first time that mutations of MYH9 result in an alteration of the composition and agonist-induced reorganization of the platelet cytoskeleton. We suggest that these abnormalities may represent the biochemical basis for the previously reported functional alterations of MYH9-RD platelets, and for the abnormal platelet formation from megakaryocytes, resulting in thrombocytopenia and giant platelets.
  Journal of Thrombosis and Haemostasis 06/2005; 3(5):1026-35. · 5.73 Impact Factor
• Article: Utility of biochemical markers in the follow-up of heart transplant recipients.

  A Balduini, C Campana, M Ceresa, E Arbustini, T Bosoni, A Serio, C Tinelli, M Viganò, G L Melzi D'Eril, L Tavazzi, R Moratti, G Merlini
  [show abstract] [hide abstract]
  ABSTRACT: Endomyocardial biopsy (EMB) is currently the standard method to diagnose acute graft rejection. However, considering the potential complications of this procedure, a noninvasive marker of rejection would be an ideal alternative or at least a helpful adjunct to posttransplant management. We measured myoglobin (Myo), creatine kinase MB mass (CK-MBm), troponin T (cTnT), serum amyloid A (SAA), and C-reactive protein (CRP) in 57 patients (mean age 37.5 years) who underwent orthotopic heart transplantation for end-stage cardiac failure between January and December 2001.Endomyocardial biopsies were performed routinely after surgery and histologically diagnosed rejection was graded according to the criteria of the International Society of Heart and Lung Transplantation. Concomittant with the biopsies, blood samples were drawn from the coronary sinus (central blood samples) and from a peripheral vein (peripheral blood samples) to assay biochemical markers. Among 149 EMB evaluated, 87 were negative (grade 0); 28 showed grade 1a rejection; 26 showed grade 1b; and 8 showed grade > 1b (2 were grade 2, 6 were grade 3a). Grades 0 and 1a were considered to be negative, while grades 1b and >1b were considered positive indicating potential acute graft rejection. cTnT, Myo, CK-MBm, SAA, and CRP levels were measured in 149 central blood samples and 149 peripheral blood samples. Myo and CK-MBm did not show significant changes. cTnT seems to be a potentially useful addition to the EMB results, while SAA and CRP showed variations with respect to EMB grade both in central and peripheral samples.
  Transplantation Proceedings 01/2004; 35(8):3075-8. · 1.00 Impact Factor
• Article: Haematological modifications after acute exposure to high altitude: possible implications for detection of recombinant erythropoietin misuse.

  M Bonfichi, A Balduini, L Arcaini, A Lorenzi, C Marseglia, L Malcovati, L Bernardi, C Passino, G Spandacini, P Feil, C Keyl, A Schneider, A Boiardi, G Bandinelli, R E Greene, C Bernasconi
  British Journal of Haematology 07/2000; 109(4):895-6. · 4.94 Impact Factor
• Article: A human factor-dependent cell line simulates cobblestone formation under human bone marrow stromal cells in vitro.

  C R Mantel, A Balduini, H E Broxmeyer
  Annals of the New York Academy of Sciences 05/1999; 872:399-401. · 3.15 Impact Factor
• Article: Comparative effects of retroviral-mediated gene transfer into primary human stromal cells of Flt3-ligand, interleukin 3 and GM-CSF on production of cord blood progenitor cells in long-term culture.

  A Balduini, S E Braun, K Cornetta, S Lyman, H E Broxmeyer
  [show abstract] [hide abstract]
  ABSTRACT: The effects of different cytokines on growth of human cord blood CD34 cells was studied by performing long-term culture (LTC) with primary human stromal cells transduced with genes for either Flt3-ligand (L) (human transmembrane, murine soluble or murine membrane-bound forms), human interleukin 3 (IL-3) or human GM-CSF. Molecular analysis of genomic DNA from transduced stromal cells using neo-specific polymerase chain reaction demonstrated gene transfer of G418-selected stromal cell populations. Enzyme-linked immunosorbent assay and biological assays of conditioned media from transduced stromal cells indicated expression and release of soluble cytokines. Numbers of both immature and more mature progenitors (colony-forming unit-granulocyte, erythroid, macrophage, megakaryocyte; CFU-GEMM, BFU-E, CFU-GM) were increased threefold compared to control in the Flt3-L (transmembrane) LTC throughout five weeks of culture. IL-3 and GM-CSF feeders increased progenitor cell output also, but these effects were significantly lower than Flt3-L feeders. The two Flt3-L isoform engineered feeders, Ex6 (soluble isoform) and 5H (membrane-bound isoform), showed a decreased effect compared to the transmembrane Flt3-L feeders and, in particular, Ex6 feeders were similar to control feeders and 5H feeders were comparable to Flt3-L feeders only in the first two weeks of LTC. These results were apparent also by limiting dilution assays that showed a higher frequency of pre-CFU in the transmembrane Flt3-L feeders compared to control and the other cytokine feeders. Exogenous addition of soluble growth factors to suspension cultures without feeder layers, while superior to stromal feeders for short-term expansion of early progenitors, were inferior to the long-term maintenance/output on stromal feeders. Pre-CFU analysis supported these data. These results may be of some significance to understanding the actions of Flt3-L on blood cell production.
  Stem Cells 02/1998; 16 Suppl 1:37-49. · 7.78 Impact Factor
• Article: Growth factors in the therapy of myelodysplasia: biological aspects.

  M Bonfichi, C Astori, E P Alessandrino, P Bernasconi, A Balduini, C Castagnola, E Brusamolino, G Pagnucco, A Canevari, P Trucco, C Bernasconi
  [show abstract] [hide abstract]
  ABSTRACT: Growth factors (GF) are reported to play an important role in the therapy of myelodisplastic syndromes (MDS). After in vitro administration a consistent group of MDS may respond to GF but the possibility of differentiation, regulation or expansion of myelodisplastic clones following GF therapy is still a question to be answered as their optimum dose and combinations. To validate if in vivo treatment with GF, may promote the regulation or the recovery of myelopoiesis and/or modify the clonality of the responses, we gave G-CSF after intensive chemotherapy in high risk MDS and acute leukemia evolving from MDS patients. According to our data the use of G-CSF after intensive chemotherapy may improve the CR rate without increase of leukemic transformation. However the answer were clonal and the remission duration remained very short so we suggest to utilize this time to perform other therapeutic strategies such as, when possible, the BMT.
  Leukemia and Lymphoma 01/1998; 26 Suppl 1:35-40. · 2.58 Impact Factor