Genomic and Proteomic Analysis of the Impact of Mitotic Quiescence on the Engraftment of Human CD34+ Cells

It is well established that in adults, long-term repopulating hematopoietic stem cells (HSC) are mitotically quiescent cells that reside in specialized bone marrow (BM) niches that maintain the dormancy of HSC. Our laboratory demonstrated that the engraftment potential of human HSC (CD34+ cells) from BM and mobilized peripheral blood (MPB) is restricted to cells in the G0 phase of cell cycle but that in the case of umbilical cord blood (UCB) -derived CD34+ cells, cell cycle status is not a determining factor in the ability of these cells to engraft and sustain hematopoiesis. We used this distinct in vivo behavior of CD34+ cells from these tissues to identify genes associated with the engraftment potential of human HSC. CD34+ cells from BM, MPB, and UCB were fractionated into G0 and G1 phases of cell cycle and subjected in parallel to microarray and proteomic analyses. A total of 484 target genes were identified to be associated with engraftment potential of HSC. System biology modeling indicated that the top four signaling pathways associated with these genes are Integrin signaling, p53 signaling, cytotoxic T lymphocyte-mediated apoptosis, and Myc mediated apoptosis signaling. Our data suggest that a continuum of functions of hematopoietic cells directly associated with cell cycle progression may play a major role in governing the engraftment potential of stem cells. While proteomic analysis identified a total of 646 proteins in analyzed samples, a very limited overlap between genomic and proteomic data was observed. These data provide a new insight into the genetic control of engraftment of human HSC from distinct tissues and suggest that mitotic quiescence may not be the requisite characteristic of engrafting stem cells, but instead may be the physiologic status conducive to the expression of genetic elements favoring engraftment

Towards a Clinically Relevant Lentiviral Transduction Protocol for Primary Human CD34+ Hematopoietic Stem/Progenitor Cells

Background: Hematopoietic stem cells (HSC), in particular mobilized peripheral blood stem cells, represent an attractive target for cell and gene therapy. Efficient gene delivery into these target cells without compromising self-renewal and multipotency is crucial for the success of gene therapy. We investigated factors involved in the ex vivo transduction of CD34 + HSCs in order to develop a clinically relevant transduction protocol for gene delivery. Specifically sought was a protocol that allows for efficient transduction with minimal ex vivo manipulation without serum or other reagents of animal origin. Methodology/Principal Findings: Using commercially available G-CSF mobilized peripheral blood (PB) CD34 + cells as the most clinically relevant target, we systematically examined factors including the use of serum, cytokine combinations, prestimulation time, multiplicity of infection (MOI), transduction duration and the use of spinoculation and/or retronectin. A self-inactivating lentiviral vector (SIN-LV) carrying enhanced green fluorescent protein (GFP) was used as the gene delivery vehicle. HSCs were monitored for transduction efficiency, surface marker expression and cellular function. We were able to demonstrate that efficient gene transduction can be achieved with minimal ex vivo manipulation while maintaining the cellular function of transduced HSCs without serum or other reagents of animal origin. Conclusions/Significance: This study helps to better define factors relevant towards developing a standard clinical protocol for the delivery of SIN-LV into CD34 + cells

Effect of rosuvastatin on outcomes in chronic haemodialysis patients – design and rationale of the AURORA study

BACKGROUND: Patients with end-stage renal disease (ESRD) are at high risk of cardiovascular events. Multiple risk factors for atherosclerosis are present in ESRD and may contribute to the increased risk of cardiovascular mortality in this population. In contrast to patients with normal renal function, the benefits of modifying lipid levels on cardiovascular outcomes in patients with ESRD on haemodialysis have yet to be confirmed in large prospective randomised trials. A study to evaluate the Use of Rosuvastatin in subjects On Regular haemodialysis: an Assessment of survival and cardiovascular events (AURORA) will be the first large-scale international trial to assess the effects of statin therapy on cardiovascular morbidity and mortality in ESRD patients on chronic haemodialysis. METHODS: More than 2,750 ESRD patients who have been receiving chronic haemodialysis treatment for at least 3 months have been randomised (1:1), irrespective of baseline lipid levels, to treatment with rosuvastatin 10 mg or placebo. The primary study endpoint is the time to a major cardiovascular event (first occurrence of cardiovascular death, non-fatal myocardial infarction or non-fatal stroke). Secondary endpoints include all-cause mortality, major cardiovascular event-free survival time, time to cardiovascular death, time to non-cardiovascular death, cardiovascular interventions, tolerability of treatment and health economic costs per life-year saved. Study medication will be given until 620 subjects have experienced a major cardiovascular event. CONCLUSION: Our hypothesis is that results from AURORA will establish the clinical efficacy and tolerability of rosuvastatin in patients with ESRD receiving chronic haemodialysis and guide the optimal management of this expanding population

Applicability of dinoflagellate cyst stratigraphy to the analyses of passive and active tectonic settings

The notion that fluctuating tectonic stress patterns within or between continental plates directly influence the development of a given sedimentary basin is a well-established concept in geotectonics. In recent years it has become increasingly understood that notably the phase of relative compressional stress build-up causes dramatic changes in basin configurations, particularly at basin margins. Detailed assessment of the timing and duration of such phases, and characterisation of the concomitant changes in sedimentary facies distribution patterns (or paleoenvironments) is vital for the better understanding of this process and the various possible underlying mechanisms. Conventionally in Mesozoic and Cenozoic basin analysis notably, the study and interpolation of ammonites and/or calcareous microfossils (planktic and benthic foraminifera, calcareous nannoplankton) play an important role in the generation of chronostratigraphically and paleoenvironmentally significant information. However, these fossil groups are frequently unsuitable for intra-basinal correlations between marginal marine and deeper marine deposits. Meanwhile, notably in hydrocarbon exploration the study and interpretation of dinoflagellate cyst assemblages has become increasingly successful in stratigraphic analysis of Mesozoic and Cenozoic basins throughout the world. Studies on Cenozoic and Recent dinoflagellate cysts published over the last decades stress their sensitivity to environmental changes, and successful application in high resolution stratigraphy (e.g., Wall et aI., 1977; papers in Head and Wrenn, 1992; Brinkhuis, 1992; Versteegh, 1995). Moreover, the organic-walled cyst producing dinoflagellates are primarily associated with neritic environments, causing their consistent occurrence in marginal marine settings. In more offshore settings, the dinoflagellate signal increasingly consists of transported elements, thus allowing detailed correlations along onshore-offshore transects. This thesis therefore concentrates on the assessment of the applicability of this relatively new biostratigraphic tool on the timing and characterisation of periods (inception) of compression. For the purpose of this study compressional phases recognised in two highly contrasting basins were selected, viz. (I) in the Early Cretaceous of the primarily carbonate dominated Dauphinois Basin (SE France), developed in a tectonically passive setting, and (2) in the Oligocene of the mainly siliciclastic-dominated Pindos Foreland Basin (Epirus, NE Greece) which developed in front of the Pindos Thrust.

Dinoflagellate cysts, glacio-eustacy and tectonics; a case study from the Eocene-Oligocene transition of the Pindos Foreland Basin (NW Greece).

In an attempt to discriminate between tectonically induced sea-level changes and glacio-eustacy, the Ekklissia and Arakthos sections (Epirus, NW Greece) are examined, applying (dinocyst) palynology, sedimentology and magnetostratigraphy. The sections, located in the Pindos Foreland Basin, both comprise the transition from pelagic limestones to hemipelagic silty clays and turbidite sandstones, reflecting the onset of flysch sedimentation as a result of the Pindos thrust activity. Despite an overall tectonic overprint, relative changes of sea level can be reconstructed, using (i) continental/marine palynomorph ratios, (ii) relative abundance of inshore and offshore dinoflagellate cysts, and (iii) taxa indicative of relatively cold and warm sea-surface temperature, that can be calibrated against the Global Polarity Time Scale (GPTS). Increased fluxes of marginal marine and continental palynomorphs coincide with colder periods on a 'third-order' scale, which thus appear to be related to glacio-eustatic trends in sea-level. The larger scale is attributed to the increasing effect of tectonics and acts on a 'second-order scale'

The effect of twin-to-twin interval on neonatal outcome of the second twin

Research into fetal development and medicin