Exploiting human CD34+ stem cell-conditioned medium for tissue repair.

Despite the progress in our understanding of genes essential for stem cell regulation and development, little is known about the factors secreted by stem cells and their effect on tissue regeneration. In particular, the factors secreted by human CD34+ cells remain to be elucidated. We have approached this challenge by performing a cytokine/growth factor microarray analysis of secreted soluble factors in medium conditioned by adherent human CD34+ cells.

MicroRNA-181a* Targets Nanog in a Subpopulation of CD34(+) Cells Isolated From Peripheral Blood.

Exploiting the properties of stem cells by microRNA (miRNA) profiling offers an attractive approach to identify new regulators of stem cell fate. Although numerous miRNA have been screened from hematopoietic stem cells (HSC), the targets corresponding to many of these miRNA have not yet been fully elucidated. By miRNA profiling in a subpopulation of CD34+ cells isolated from peripheral blood, we have identified eight clusters of miRNA that were differentially expressed.

Protein segregation between dividing hematopoietic progenitor cells in the determination of the symmetry/asymmetry of cell division.

In the present study, we investigated how the symmetry/asymmetry of cell division in mitotic CD34(+) cells can be evaluated by determining the plane of cell division and the potential distribution of proteins between daughter cells. The orientation of the mitotic spindle is dependent upon the positioning of the centrosomes, which determine the plane of cell division and the sharing of proteins. If the functions of unequally shared proteins are relevant to the kinetics of cell division, they could determine whether the daughter cells undergo self-renewal or differentiation.

Suppression of erythropoiesis in patients with chronic heart failure and anaemia of unknown origin: evidence of an immune basis.

Aims: Anaemia is a prevalent and adverse comorbidity in chronic heart failure (CHF) but its origins are frequently elusive. Diffuse inflammation is also prominent in CHF and a potent inhibitor of erythrocyte production. We tested the hypothesis that unexplained anaemia in CHF might be subsequent to diminished erythropoiesis as a result of an immune-mediated suppression of erythroid colony formation.

Erythropoietin resistance contributes to anaemia in chronic heart failure and relates to aberrant JAK-STAT signal transduction.

Background: Chronic heart failure (CHF) patients are frequently anaemic despite elevated endogenous erythropoietin (Epo) levels. We tested the hypothesis that this might be due to Epo resistance and investigated whether any defects apparent were due to Epo receptor (EpoR) downregulation and/or impaired Epo-induced signal transduction.

Methods: We studied 28 CHF patients (age 64 ± 10 yrs, LVEF 29 ± 9%, 89% male) and 12 healthy controls (65 ± 11 yrs, 75% male).

Antibody arrays identify protein-protein interactions in chronic myeloid leukaemia.

Multiprotein complex formation with p210(BCR-ABL1) is likely to play a major role in determining cellular abnormalities in chronic myeloid leukaemia (CML). Although many p210(BCR-ABL1) binding partners have been identified, it is likely that many have not. We evaluated the use of co-immunoprecipitation and antibody arrays and found that this approach is capable of identifying new p210(BCR-ABL1) binding partners, and may contribute to the search for new therapeutic targets in CML.

Influence of PI-3K/Akt pathway on Wnt signalling in regulating myeloid progenitor cell proliferation. Evidence for a role of autocrine/paracrine Wnt regulation.

The regulation of myeloid progenitor cell (granulocyte-macrophage colony-forming units, CFU-GM) proliferation/differentiation by the Wnt and phosphatidylinositol-3 kinase (PI-3K) pathways was investigated using a colony-replating assay. The PI-3K pathway promoted differentiation of interleukin-3 (IL-3)-stimulated myelopoiesis via Akt, because inhibition of the PI-3K/Akt pathway with LY294002 or SH-5 increased proliferation. The involvement of canonical and non-canonical Wnt pathways was investigated using Wnt3a and Wnt5a respectively.

Abnormal centrosome-centriole cycle in chronic myeloid leukaemia?

Abnormal numbers, structures and functions of centrosomes in chronic myeloid leukaemia (CML) may influence cell proliferation and genomic instability, which are features of the disease. Centrosomes are regulators of mitotic spindle orientation and can act as scaffolds for centrosome-associated regulators of the cell cycle. This study showed, for the first time, that p210(BCR-ABL1) and p145(ABL1) are both centrosome-associated proteins, as demonstrated by co-immunoprecipitation with the pericentriolar protein, pericentrin.

Autologous infusion of expanded mobilized adult bone marrow-derived CD34+ cells into patients with alcoholic liver cirrhosis.

Objectives: Recent advances in regenerative medicine, including hematopoietic stem cell (HSC) transplantation, have brought hope for patients with severe alcoholic liver cirrhosis (ALC). The aim of this study was to assess the safety and efficacy of administering autologous expanded mobilized adult progenitor CD34+ cells into the hepatic artery of ALC patients and the potential improvement in the liver function.

Methods: Nine patients with biopsy-proven ALC, who had abstained from alcohol for at least 6 months, were recruited into the study.

Stem cells for regenerative medicine--biological attributes and clinical application.

The place of stem cells in the therapeutic armamentarium is currently subject to intense scrutiny. One issue is the nature of the stem cells themselves, and their relationship to better-known stem cells, such as hematopoietic stem cells. Here it is argued that the stem cells we propose for therapeutic application may be more closely related to embryonic cells than are hematopoietic stem cells.

Strontium can increase some osteoblasts without increasing hematopoietic stem cells.

Osteoblasts are a key component in the regulation of the hematopoietic stem cell (HSC) niche. Manipulating osteoblast numbers results in a parallel change in HSC numbers. We tested the activity of strontium (Sr), a bone anabolic agent that enhances osteoblast function and inhibits osteoclast activity, on hematopoiesis.

Potential of CD34 in the regulation of symmetrical and asymmetrical divisions by hematopoietic progenitor cells.

The control of symmetric and asymmetric division in the hematopoietic stem/progenitor cell population is critically important for the regulation of blood cell production. Asymmetric divisions depend on cell polarization, which may be conferred by location and/or interaction with neighboring cells. In this study, we sought evidence for polarization in CD34+ cells, which interact by binding to one another.

A novel HSV-1 virus, JS1/34.5-/47-, purges contaminating breast cancer cells from bone marrow.

Purpose: Oncolytic herpes simplex virus type 1 (HSV-1) vectors show considerable promise as agents for cancer therapy. We have developed a novel recombinant HSV-1 virus (JS1/34.5-/47-) for purging of occult breast cancer cells from bone marrow of patients.

Detection in primary chronic myeloid leukaemia cells of p210BCR-ABL1 in complexes with adaptor proteins CBL, CRKL, and GRB2.

Chronic myeloid leukemia (CML) arises as a consequence of the expression of a chimeric fusion protein, p210BCR-ABL1, which is localized to the cytoplasm and has constitutive protein tyrosine kinase activity. Extensive publications report that p210BCR-ABL1 complexed with multiple cytoplasmic proteins can modulate several cell signaling pathways. However, while altered signaling states can be demonstrated in primary CML material, most of the reported analytical work on complexed proteins has been done in cell lines expressing p210BCR-ABL1.

Adult bone marrow-derived stem cells and the injured heart: just the beginning?

Coronary artery disease leading to ischaemia of the human myocardium is one of the chief causes of morbidity and mortality in the western world. Cellular transplantation has recently been proposed as a novel alternative treatment modality. Adult bone marrow-derived autologous cells are one of the key cell types under investigation in both the experimental and clinical setting.

Chronic myeloid leukaemia: stem cell derived but progenitor cell driven.

The biology of CML (chronic myeloid leukaemia) has been extensively investigated as the disease is a paradigm of neoplasms induced when a translocation results in expression of a novel fusion protein, in this instance p210(BCR-ABL). Although CML manifests itself principally as unregulated expansion of the myeloid lineage, the lesion is present in the stem cell population and it has long been assumed that disregulated stem cell kinetics must underlie the basic pathology of the disease. In this review, we present evidence that, in normal haemopoiesis, less primitive precursor cells retain considerable flexibility in their capacity to undergo self-renewal, allowing them to maintain lineage-specific homoeostasis without inflicting proliferative stress upon the stem cell population.

Targeting primary human leukaemia cells with RNA interference: Bcr-Abl targeting inhibits myeloid progenitor self-renewal in chronic myeloid leukaemia cells.

We have investigated functional outcome of challenging primary chronic myeloid leukaemia (CML) cells with Bcr-Abl fusion sequence-directed RNA interference (RNAi). We targeted the Bcr-Abl b3a2 variant, by RNAi, in primary chronic phase CML cells, and detected strikingly reduced proliferation of myeloid precursor cells expressing this variant. Lack of an effect in cells expressing a distinct Bcr-Abl variant confirmed the specificity of the response.

Growth factor displayed on the surface of retroviral particles without manipulation of envelope proteins is biologically active and can enhance transduction.

Background: The therapeutic potential of retroviruses can be significantly enhanced by display of specific molecules on the retroviral surface. This has been conventionally achieved by the manipulation of retroviral envelope proteins. In this report we have tested whether the natural budding mechanism of the retrovirus could be exploited to incorporate a specific molecule into the retroviral surface.

Targeted retroviral transduction of c-kit+ hematopoietic cells using novel ligand display technology.

Gene therapy for a wide variety of disorders would be greatly enhanced by the development of vectors that could be targeted for gene delivery to specific populations of cells. We describe here high-efficiency targeted transduction based on a novel targeting strategy that exploits the ability of retroviruses to incorporate host cell proteins into the surface of the viral particle as they bud through the plasma membrane. Ecotropic retroviral particles produced in cells engineered to express the membrane-bound form of stem cell factor (mbSCF) transduce both human cell lines and primary cells with high efficiency in a strictly c-kit (SCF receptor)-dependent fashion.

Opposing effects of PI3 kinase pathway activation on human myeloid and erythroid progenitor cell proliferation and differentiation in vitro.

Objective: To investigate 1) the effects of lineage-specific cytokines (G-CSF and EPO) combined with ligands for different classes of cytokine receptors (common beta chain, gp130, and tyrosine kinase) on proliferation by human myeloid and erythroid progenitor cells; and 2) the signal transduction pathways associated with combinatorial cytokine actions.

Patients And Methods: CFU-GM and BFU-E were cloned in vitro. Secondary colony formation by replated CFU-GM and subcolony formation by BFU-E provided measures of progenitor cell proliferation.

Clinical heterogeneity in chronic myeloid leukaemia reflecting biological diversity in normal persons.

The molecular basis of chronic myeloid leukaemia (CML) is well defined and highly consistent, yet prognosis varies considerably. This could reflect the biological diversity occurring in normal populations. We used a colony replating assay to measure the proliferative capacity of progenitor cells from 211 CML patients and 86 normal persons.

Progenitor cells divide symmetrically to generate new colony-forming cells and clonal heterogeneity.

Self-renewal is the most fundamental property of haemopoietic stem and progenitor cells. However, because of the need to produce differentiated cells, not all cell divisions involve self-renewal. We have used a colony replating assay to follow the fates of individual haemopoietic progenitor cell clones.