Charalampos Pontikoglou

Specific Lineage‐Priming of Bone Marrow Mesenchymal Stem Cells Provides the Molecular Framework for Their Plasticity

Lineage‐priming is a molecular model of stem cell (SC) differentiation in which proliferating SCs express a subset of genes associated to the differentiation pathways to which they can commit. This concept has been developed for hematopoietic SCs, but has been poorly studied for other SC populations. Because the differentiation potential of human bone marrow mesenchymal stem cells (BM MSCs) remains controversial, we have explored the theory of lineage‐priming applied to these cells. We show that proliferating primary layers and clones of BM MSCs have precise priming to the osteoblastic (O), chondrocytic (C), adipocytic (A), and the vascular smooth muscle (V) lineages, but not to skeletal muscle, cardiac muscle, hematopoietic, hepatocytic, or neural lineages. Priming was shown both at the mRNA (300 transcripts were evaluated) and the protein level. In particular, the master transactivator proteins PPARG, RUNX2, and SOX9 were coexpressed before differentiation induction in all cells from incipient clones. We further show that MSCs cultured in the presence of inducers differentiate into the lineages for which they are primed. Our data point out to a number of signaling pathways that might be activated in proliferating MSCs and would be responsible for the differentiation and proliferation potential of these cells. Our results extend the notion of lineage‐priming and provide the molecular framework for inter‐A, ‐O, ‐C, ‐V plasticity of BM MSCs. Our data highlight the use of BM MSCs for the cell therapy of skeletal or vascular disorders, but provide a word of caution about their use in other clinical indications. Stem Cells 2009;27:1142–1151

Mesenchymal stem cells derived from Wharton's Jelly of the umbilical cord: biological properties and emerging clinical applications.

In recent years there seems to be an unbounded interest concerning mesenchymal stem cells (MSCs). This is mainly attributed to their exciting characteristics including long-term ex vivo proliferation, multilineage potential and immunomodulatory properties. In this regard MSCs emerge as attractive candidates for various therapeutic applications. MSCs were originally isolated from the bone marrow (BM) and this population is still considered as the gold standard for MSC applications. Nevertheless the BM has several limitations as source of MSCs, including MSC low frequency in this compartment, the painful isolation procedure and the decline in MSC characteristics with donors age. Thus, there is accumulating interest in identifying alternative sources for MSCs. To this end MSCs obtained from the Whartons Jelly (WJ) of umbilical cords (UC) have gained much attention over the last years since they can be easily isolated, without any ethical concerns, from a tissue which is discarded after birth. Furthermore WJ-derived MSCs represent a more primitive population than their adult counterparts, opening new perspectives for cell-based therapies. In this review we will at first give an overview of the biology of WJ-derived UC-MSCs. Then their potential application for the treatment of cancer and immune mediated disorders, such graft versus host disease (GVHD) and systemic lupus erythematosus (SLE) will be discussed, and finally their putative role as feeder layer for ex vivo hematopoietic stem cell (HSC) expansion will be pointed out.

Bone Marrow Mesenchymal Stem Cells: Biological Properties and Their Role in Hematopoiesis and Hematopoietic Stem Cell Transplantation

Mesenchymal stem cells (MSCs) are multipotent adult stem cells that are present in practically all tissues as a specialized population of mural cells/pericytes that lie on the abluminal side of blood vessels. Originally identified within the bone marrow (BM) stroma, not only do they provide microenvironmental support for hematopoietic stem cells (HSCs), but can also differentiate into various mesodermal lineages. MSCs can easily be isolated from the BM and subsequently expand in vitro and in addition they exhibit intriguing immunomodulatory properties, thereby emerging as attractive candidates for various therapeutic applications. This review addresses the concept of BM MSCs via a hematologist’s point of view. In this context it discusses the stem cell properties that have been attributed to BM MSCs, as compared to those of the prototypic hematopoietic stem cell model and then gives a brief overview of the in vitro and vivo features of the former, emphasizing on their immunoregulatory properties and their hematopoiesis-supporting role. In addition, the qualitative and quantitative characteristics of BM MSCs within the context of a defective microenvironment, such as the one characterizing Myelodysplastic Syndromes are described and the potential involvement of these cells in the pathophysiology of the disease is discussed. Finally, emerging clinical applications of BM MSCs in the field of hematopoietic stem cell transplantation are reviewed and potential hazards from MSC use are outlined.

Effect of lenalidomide therapy on hematopoiesis of patients with myelodysplastic syndrome associated with chromosome 5q deletion

Background Lenalidomide improves erythropoiesis in patients with low/intermediate-1 risk myelodysplastic syndrome and interstitial deletion of the long arm of chromosome 5 [del(5q)]. The aim of this study was to explore the effect of lenalidomide treatment on the reserves and functional characteristics of bone marrow hematopoietic progenitor/precursor cells, bone marrow stromal cells and peripheral blood lymphocytes in patients with low/intermediate-1 risk myelodysplastic syndrome with del(5q). Design and Methods We evaluated the number and clonogenic potential of bone marrow erythroid/myeloid/megakaryocytic progenitor cells using clonogenic assays, the apoptotic characteristics and adhesion molecule expression of CD34+ cells by flow cytometry, the hematopoiesis-supporting capacity of bone marrow stromal cells using long-term bone marrow cultures and the number and activation status of peripheral blood lymphocytes in ten patients with low/intermediate-1 risk myelodysplastic syndrome with del(5q) receiving lenalidomide. Results Compared to baseline, lenalidomide treatment significantly decreased the proportion of bone marrow CD34+ cells, increased the proportion of CD36+/GlycoA+ and CD36−/GlycoA+ erythroid cells and the percentage of apoptotic cells within these cell compartments. Treatment significantly improved the clonogenic potential of bone marrow erythroid, myeloid, megakaryocytic colony-forming cells and increased the proportion of CD34+ cells expressing the adhesion molecules CD11a, CD49d, CD54, CXCR4 and the SLAM antigen CD48. The hematopoiesis-supporting capacity of bone marrow stroma improved significantly following treatment, as demonstrated by the number of colony-forming cells and the level of stromal-derived factor-1α and intercellular adhesion molecule-1 in long-term bone marrow culture supernatants. Lenalidomide treatment also increased the proportion of activated peripheral blood T lymphocytes. Conclusions The beneficial effect of lenalidomide in patients with lower risk myelodysplastic syndrome with del(5q) is associated with significant increases in the proportion of bone marrow erythroid precursor cells and in the frequency of clonogenic progenitor cells, a substantial improvement in the hematopoiesis-supporting potential of bone marrow stroma and significant alterations in the adhesion profile of bone marrow CD34+ cells.

Normal bone marrow hematopoietic stem cell reserves and normal stromal cell function support the use of autologous stem cell transplantation in patients with multiple sclerosis.

Summary:Bone marrow (BM) stem cell reserves and function and stromal cell hematopoiesis supporting capacity were evaluated in 15 patients with multiple sclerosis (MS) and 61 normal controls using flow cytometry, clonogenic assays, long-term BM cultures (LTBMCs) and enzyme-linked immunosorbent assays. MS patients displayed normal CD34+ cell numbers but a low frequency of colony-forming cells (CFCs) in both BM mononuclear and purified CD34+ cell fractions, compared to controls. Patients had increased proportions of activated BM CD3+/HLA-DR+ and CD3+/CD38+ T cells that correlated inversely with CFC numbers. Patient BM CD3+ T cells inhibited colony formation by normal CD34+ cells and patient CFC numbers increased significantly following immunomagnetic removal of T cells from BMMCs, suggesting that activated T cells may be involved in the defective clonogenic potential of hematopoietic progenitors. Patient BM stromal cells displayed normal hematopoiesis supporting capacity indicated by the CFC number in the nonadherent cell fraction of LTBMCs recharged with normal CD34+ cells. Culture supernatants displayed normal stromal derived factor-1 and stem cell factor/kit ligand but increased flt-3 ligand levels. These findings provide support for the use of autologous stem cell transplantation in MS patients. The low clonogenic potential of BM hematopoietic progenitors probably reflects the presence of activated T cells rather than an intrinsic defect.

Bone regeneration: the stem/progenitor cells point of view

•  Introduction •  Mesenchymal stem cells as skeletal stem cells ‐  Cultured or expanded MSCs ‐  Native MSCs ‐  The neural crest origin of MSCs •  Vascularization by endothelial progenitor cells as prerequisite process before bone repair ‐  Vascularization and bone healing ‐  Endothelial progenitor cells •  Conclusion

Human bone marrow native mesenchymal stem cells.

The adult bone marrow (BM) has been generally considered to be composed of hematopoietic tissue and the associated supporting stroma. There is accumulating evidence that a subset of multipotential cells exists within the latter compartment, commonly referred to as mesenchymal stem cells (MSCs). These cells can be easily expanded ex vivo and induced to differentiate into skeletal connective tissues, thereby emerging as attractive candidates for various therapeutic applications. Despite the extensive in vitro characterization of MSCs, at present, little is known about their in vivo properties or anatomical location. Here we review the efficacy of the different surface markers used to isolate BM MSCs and highlight current data suggesting that BM MSCs in situ are associated with vascular sinus walls and probably belong to the family of vascular smooth muscle cells.

Lymphocyte subpopulation imbalances, bone marrow hematopoiesis and histopathology in rituximab-treated lymphoma patients with late-onset neutropenia

Lymphocyte subpopulation imbalances, bone marrow hematopoiesis and histopathology in rituximab-treated lymphoma patients with late-onset neutropenia

Idiosyncratic Drug-Induced Agranulocytosis: The Paradigm of Deferiprone

Non chemotherapy drug-induced agranulocytosis is considered a potentially life-threatening idiosyncratic blood dyscrasia, thought to result from a partly elucidated immune and/or toxic damage on myelopoiesis, due to a multitude of drugs. Offending agents include clozapine, ticlopidine, antithyroid compounds, dipyrone, sulfasalzine, trimethropim/sulfomethoxazole, carmabazepine, and to a lesser extent, deferiprone (L1) and probably rituximab. Suspected drugs should be immediately stopped and, in symptomatic patients, once appropriate cultures have been obtained, broad-spectrum antibiotic treatment should be administered. Hematopoietic growth factors may be considered, specifically in patients with poor prognostic factors. Due to improved intensive care treatment and alertness of physicians the case fatality of the disorder has recently been decreased to 5%.

Pathophysiologic mechanisms, clinical features and treatment of idiopathic neutropenia

The term idiopathic neutropenia describes a benign disorder of granulopoiesis characterized by the unexplained reduction in the absolute neutrophil count below the lower limit of the normal range for a prolonged period. Recent studies have provided evidence that this neutropenic condition comprises two distinct disease entities on the basis of the underlying pathogenetic mechanisms: first, the primary autoimmune neutropenia mediated by autoantibodies against mature neutrophils and/or their bone marrow progenitor/precursor cells; and second, the previously named chronic idiopathic neutropenia, that might now be called chronic immunologic neutropenia, characterized by T-cell- and cytokine-mediated suppression of granulopoiesis. Despite the differences in the bone marrow granulocytic progenitor cell reserves that actually reflect the differences in the implicated pathophysiologic mechanisms, both disease entities usually display an uncomplicated clinical course with minimal symptoms. Treatment decisions should be individualized on the basis of patients’ clinical course and the indicated therapies are analyzed in this review. The clinical and laboratory data characterizing these neutropenic conditions and the available in vitro data that have led to remarkable progress in the understanding of the pathophysiology of both disorders are also summarized.