Common molecular pathways involved in human CD133+/CD34+ progenitor cell expansion and cancer

<p>Abstract</p> <p>Background</p> <p>Uncovering the molecular mechanism underlying expansion of hematopoietic stem and progenitor cells is critical to extend current therapeutic applications and to understand how its deregulation relates to leukemia. The characterization of genes commonly relevant to stem/progenitor cell expansion and tumor development should facilitate the identification of novel therapeutic targets in cancer.</p> <p>Methods</p> <p>CD34+/CD133+ progenitor cells were purified from human umbilical cord blood and expanded <it>in vitro</it>. Correlated molecular changes were analyzed by gene expression profiling using microarrays covering up to 55,000 transcripts. Genes regulated during progenitor cell expansion were identified and functionally classified. Aberrant expression of such genes in cancer was indicated by <it>in silico </it>SAGE. Differential expression of selected genes was assessed by real-time PCR in hematopoietic cells from chronic myeloid leukemia patients and healthy individuals.</p> <p>Results</p> <p>Several genes and signaling pathways not previously associated with <it>ex vivo </it>expansion of CD133+/CD34+ cells were identified, most of which associated with cancer. Regulation of MEK/ERK and Hedgehog signaling genes in addition to numerous proto-oncogenes was detected during conditions of enhanced progenitor cell expansion. Quantitative real-time PCR analysis confirmed down-regulation of several newly described cancer-associated genes in CD133+/CD34+ cells, including <it>DOCK4 </it>and <it>SPARCL1 </it>tumor suppressors, and parallel results were verified when comparing their expression in cells from chronic myeloid leukemia patients</p> <p>Conclusion</p> <p>Our findings reveal potential molecular targets for oncogenic transformation in CD133+/CD34+ cells and strengthen the link between deregulation of stem/progenitor cell expansion and the malignant process.</p

Identification of Novel Immunoregulatory Molecules in Human Thymic Regulatory CD4+CD25+ T Cells by Phage Display

Thymic CD4+CD25+ cells play an important role in immune regulation and are continuously developed in the thymus as an independent lineage. How these cells are generated, what are their multiple pathways of suppressive activity and which are their specific markers are questions that remain unanswered. To identify molecules involved in the function and development of human CD4+CD25+ T regulatory cells we targeted thymic CD4+CD25+ cells by peptide phage display. A phage library containing random peptides was screened ex vivo for binding to human thymic CD4+CD25+ T cells. After four rounds of selection on CD4+CD25+ enriched populations of thymocytes, we sequenced several phage displayed peptides and selected one with identity to the Vitamin D Receptor (VDR). We confirmed the binding of the VDR phage to active Vitamin D in vitro, as well as the higher expression of VDR in CD4+CD25+ cells. We suggest that differential expression of VDR on natural Tregs may be related to the relevance of Vitamin D in function and ontogeny of these cells

Predictors of high-quality cord blood units

BACKGROUNDAnalysis of umbilical cord blood (UCB) transplants shows a correlation between engraftment and total number of infused cells. Thus, it is worth evaluating what maternal and neonatal characteristics and collection techniques may affect the quality of UCB units. STUDY DESIGN AND METHODSA cross-sectional study was performed with 7897 donors sequentially selected in three health care institutions in Brazil from October 2004 to March 2012, in which both quantitative and qualitative approaches were applied. All donors were considered suitable for cord blood collection. RESULTSThe maternal and neonatal characteristics and techniques of collection that influenced the total number of nucleated cells (TNCsp<0.001) were type of delivery, newborn weight and sex, and institution of UCB collection. The TNC count was associated with gestational age (p=0.008), type of delivery (p<0.001), newborn sex (p<0.001), newborn weight (p<0.001), and UCB collection technique (p=0.003). Center B presented the largest number of nucleated cells in its results (p<0.001), followed by Center A (p=0.001). Other characteristics, such as maternal age, were analyzed but were not relevant for the nucleated cell number. CONCLUSIONThis study provides elements for a model that allows an efficient selection of UCB donors, prioritizing candidates who have a better chance to lead to an optimized use of cord blood cells units.Univ Fed Sao Paulo UNIFESP, Escola Paulista Enfermagem, Sao Paulo, BrazilInst Nacl Cardiol, Rio De Janeiro, BrazilInst Israelita Ensino & Pesquisa Albert Einstein, Sao Paulo, BrazilHosp Israelita Albert Einstein, Dept Hemoterapia, Sao Paulo, BrazilUniv Fed Sao Paulo UNIFESP, Escola Paulista Enfermagem, Sao Paulo, BrazilWeb of Scienc

Mesenchymal stem cell-like properties of CD133+ glioblastoma initiating cells

Glioblastoma is composed of dividing tumor cells, stromal cells and tumor initiating CD133+ cells. Recent reports have discussed the origin of the glioblastoma CD133+ cells and their function in the tumor microenvironment. The present work sought to investigate the multipotent and mesenchymal properties of primary highly purified human CD133+ glioblastoma-initiating cells. To accomplish this aim, we used the following approaches: i) generation of tumor subspheres of CD133+ selected cells from primary cell cultures of glioblastoma; ii) analysis of the expression of pluripotency stem cell markers and mesenchymal stem cell (MSC) markers in the CD133+ glioblastoma-initiating cells; iii) side-by-side ultrastructural characterization of the CD133+ glioblastoma cells, MSC and CD133+ hematopoietic stem cells isolated from human umbilical cord blood (UCB); iv) assessment of adipogenic differentiation of CD133+ glioblastoma cells to test their MSC-like in vitro differentiation ability; and v) use of an orthotopic glioblastoma xenograft model in the absence of immune suppression. We found that the CD133+ glioblastoma cells expressed both the pluripotency stem cell markers (Nanog, Mush-1 and SSEA-3) and MSC markers. In addition, the CD133+ cells were able to differentiate into adipocyte-like cells. Transmission electron microscopy (TEM) demonstrated that the CD133+ glioblastoma-initiating cells had ultrastructural features similar to those of undifferentiated MSCs. In addition, when administered in vivo to non-immunocompromised animals, the CD133+ cells were also able to mimic the phenotype of the original patient’s tumor. In summary, we showed that the CD133+ glioblastoma cells express molecular signatures of MSCs, neural stem cells and pluripotent stem cells, thus possibly enabling differentiation into both neural and mesodermal cell types

Characterization of Adherent Umbilical Cord Blood Stromal Cells Regarding Passage, Cell Number, and Nano-biomarking Utilization

Adherent umbilical cord blood stromal cells (AUCBSCs) are multipotent cells with differentiation capacities. Therefore, these cells have been investigated for their potential in cell-based therapies. Quantum Dots (QDs) are an alternative to organic dyes and fluorescent proteins because of their long-term photostability. In this study we determined the effects of the cell passage on AUCBSCs morphology, phenotype, and differentiation potential. QDs labeled AUCBSCs in the fourth cell passage were differentiated in the three mesodermal lineages and were evaluated using cytochemical methods and transmission electron microscopy (TEM). Gene and protein expression of the AUCBSCs immunophenotypic markers were also evaluated in the labeled cells by real-time quantitative PCR and flow cytometry. In this study we were able to define the best cellular passage to work with AUCBSCs and we also demonstrated that the use of fluorescent QDs can be an efficient nano-biotechnological tool in differentiation studies because labeled cells do not have their characteristics compromised.Instituto de Ensino e Pesquisa Albert EinsteinFundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP)Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq

Expression levels of , , , , and genes in healthy individuals (control) and chronic myeloid leukemia patients

<p><b>Copyright information:</b></p><p>Taken from "Common molecular pathways involved in human CD133+/CD34+ progenitor cell expansion and cancer"</p><p>http://www.cancerci.com/content/7/1/11</p><p>Cancer Cell International 2007;7():11-11.</p><p>Published online 8 Jun 2007</p><p>PMCID:PMC1904434.</p><p></p> Total RNA was extracted from purified mononuclear cells from peripheral blood samples. Transcript levels were quantified by real-time PCR and results are given as normalized gene expression. Horizontal bars indicate median expression values. Statistical significance: < 0.001 for ; < 0.01 for , , and ; < 0.05 for and

Expansion of stem and progenitor cells from UCB cultivated in the presence or absence of estradiol

<p><b>Copyright information:</b></p><p>Taken from "Common molecular pathways involved in human CD133+/CD34+ progenitor cell expansion and cancer"</p><p>Cancer Cell International 2007;7():11-11.</p><p>Published online 8 Jun 2007</p><p>PMCID:PMC1904434.</p><p></p> (A) Fold increment in CD133+/CD34+, CD133-/CD34+, and CD133+/CD34- cell number after 7, 14, and 21 days in culture. (B) Flow cytometry analysis showing percentage of mononucleated cells expressing CD133 prior to and after culturing for 7 days. Positive and isotype controls are provided in supplemental figure S2. (C) Functional clonogenic assay based on the frequency of colony forming units (CFU). Data correspond to fold expansion of myeloid and endothelial progenitors at culture day 7. GF = basal medium supplemented with growth factors only (SCF, IL3, IL6, and Flt3-ligand); E2 = medium supplemented with growth factors plus 10 nM estradiol. Statistical significance for GF vs. E2 comparisons: * < 0.01, **< 0.001