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Pozzobon, Michela (2008) Isolation, expansion and differentiation of human bone marrow CD133+ cells: plasticity and cardiac regeneration. [Ph.D. thesis]

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Abstract (english)

The use of adult stem cells to regenerate damaged tissue circumvents the moral and technical issues associated with the use of those from an embryonic source.
Mesenchymal stem cells (MSC) can be isolated from a variety of tissues, most commonly from the bone marrow (BM), and, although they represent a very small percentage of these cells, are easily expandable. Recently, the use of MSC has provided clinical benefit to patients with osteogenesis imperfecta, graft-versushost disease and myocardial infarction.
Beside these stem cells, it is known that the bone marrow CD133+ cells play an important role in the hematopoietic compartment. The cells indeed can take part to vascular reconstitution when become endothelial cells (EC), to skeletal muscle fiber regeneration when switch in muscle precursors, and to cardiomyocytes phenotypic conversion when differentiate in cardiomyocytes like cells. While the role on hematopoiesis and vasculogenesis of the selected cells is well established, their ability to differentiate along multiple non-EC lineages has not yet been fully elucidated.
The goal of this study is to assert whether human CD133+-BM derived cells, compared with MSCs, are able to differentiate in vitro besides to blood cells, to cell lineages pertinent to the mesoderm germ layers.
To this end CD133+ cells have been isolated using a clinically approved methodology and their differentiation potential compared to that of hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs) obtained from the same BM samples.
In adopted culture conditions, CD133 expression was consistently decreased after passage 2, as well as the expression of the stemnesss markers c-kit and OCT4, whereas expression of Stage Specific Embryonic Antigen 4 (SSEA4) remain consistent on all different conditions. Expanded CD133 were also positive for HLA-ABC, but negative for HLA-DR in accordance to what has been previously reported for MSCs. Moreover they were able to differentiate into adipocytes, myoblasts, endothelial cells, osteocytes, cardiomyocytes and neuronal precursor cells.
The results of this study fully support the notion of a wide range differentiation potential of CD133+-BM derived cells, encompassing not only mesodermal but also ectodermic (neurogenic) cell lineages.
CD133 antigen could be potentially used to select a cell population with similar characteristics to the MSCs ones; the obtained results remark the great potential of CD133+ cells from BM, and support the existence of a broadly multipotent/pluripotent cell that persists in the adult, and justify the upcoming interest for possible therapeutic applications.
In vivo, using a tissue engineering approach, it has been investigated the possibility that a biodegradable and biocompatible collagen polymer, called cardiac patch, applied on the infarction area of nude rat, can give hospitality to stem cells and deliver them to improve cardiac functionality. Cells have been injected into the patch ( model I) and systemically (model II); smooth muscle actina and Von Willebrand antibodies detected many new vases in patch and cryoinjury area (as already found by the research group); about 2% of cells survived and showed good mobility into the collagen patch; improved functionality was detected compared with control animals however no cell engraftment was seen after four week from cell injection either locally or systemically.
Summarizing, the two different in vivo models aimed at define both the patch strength after cell injection in model I and the patch trophic effect in model II.
Therefore to address these points it will be mandatory to analyze bcl-2 expression on cells treated and un-treated animals, with and without patch, to verify whether the tissue engineering approach with this specific polymer, could enhance the paracrine effect of human CD133+, or suggest a change of the polymer to ameliorate cell survival.

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EPrint type:Ph.D. thesis
Tutor:Gamba, Piergiorgio
Data di deposito della tesi:2008
Anno di Pubblicazione:2008
Key Words:CD133, human bone marrow stem cells, cardiac tissue engineering
Settori scientifico-disciplinari MIUR:Area 06 - Scienze mediche > MED/11 Malattie dell'apparato cardiovascolare
Struttura di riferimento:Dipartimenti > pre 2012 - Dipartimento di Scienze Farmaceutiche
Codice ID:223
Depositato il:28 Oct 2008
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