Hilal Gul

Valproic Acid Stimulates Proliferation and Self-renewal of Hematopoietic Stem Cells

Histone deacetylase inhibitors have attracted considerable attention because of their ability to overcome the differentiation block in leukemic blasts, an effect achieved either alone or in combination with differentiating agents, such as all-trans retinoic acid. We have previously reported favorable effects of the potent histone deacetylase inhibitor valproic acid in combination with all-trans retinoic acid in patients with advanced acute myeloid leukemia leading to blast cell reduction and improvement of hemoglobin. These effects were accompanied by hypergranulocytosis most likely due to an enhancement of nonleukemic myelopoiesis and the suppression of malignant hematopoiesis rather than enforced differentiation of the leukemic cells. These data prompted us to investigate the effect of valproic acid on normal hematopoietic stem cells (HSC). Here we show that valproic acid increases both proliferation and self-renewal of HSC. It accelerates cell cycle progression of HSC accompanied by a down-regulation of p21(cip-1/waf-1). Furthermore, valproic acid inhibits GSK3beta by phosphorylation on Ser9 accompanied by an activation of the Wnt signaling pathway as well as by an up-regulation of HoxB4, a target gene of Wnt signaling. Both are known to directly stimulate the proliferation of HSC and to expand the HSC pool. In summary, we here show that valproic acid, known to induce differentiation or apoptosis in leukemic blasts, stimulates the proliferation of normal HSC, an effect with a potential effect on its future role in the treatment of acute myeloid leukemia.

Magnetic carbon nanotube labelling for haematopoietic stem/progenitor cell tracking

Haematopoietic stem and progenitor cell (HSPC) research has significantly contributed to the understanding and harnessing of haematopoiesis for regenerative medicine. However, the methodology for real-time tracking HSPC in vivo is still lacking, which seriously restricts the progress of research. Recently, magnetic carbon nanotubes (mCNT) have generated great excitement because they have been successfully used as vehicles to deliver a lot of biomolecules into various cells. There is, however, no report about mCNT being used for tracking HSPC. In this paper, we investigated the uptake efficiency of fluorescein-isothiocyanate-labelled mCNT (FITC-mCNT) into HSPC and their effect on the cytotoxicity and differentiation of HSPC. We found that cellular uptake of FITC-mCNT was concentration-and time-dependent. The uptake of FITC-mCNT into HSPC reached up to 100% with the highest mean fluorescence (MF). More importantly, efficient FITC-mCNT uptake has no adverse effect on the cell viability, cytotoxicity and differentiation of HSPC as confirmed by colony-forming unit assay (CFU). In conclusion, the results reported here suggest the further tailoring of mCNT for their use in HSPC labelling/tracking in vivo or gene delivery into HSPC.

Valproic acid exerts differential effects on CXCR4 expression in leukemic cells.

We recently reported that the histone deacetylase inhibitor, valproic acid (VPA), increases CXCR4 receptor expression and function in cord blood hematopoietic stem/progenitor cells (HSPC) and the immature, highly CD34-positive AML cell lines KG-1a and KG-1. In this study, we investigated whether VPA influences CXCR4 in CD34-negative AML cell lines (promyelocytic HL-60 and monocytic THP-1), as well as both CD34-positive and CD34-negative primary AML cells. We found that VPA (i) diminishes CXCR4 expression and chemotaxis in HL-60 cells and in the CD34-negative subtypes of primary AML cells and (ii) increases CXCR4 expression and function in the highly CD34-positive subtypes of primary AML cells. Hence, we suggest that VPA exerts different effects on CXCR4 depending on cell maturation status, and this novel finding may have important implications for AML therapy.

Light scattering characterization of single biological cells in a microfluidic cytometer

The characterization of single biological cells in a microfluidic flow by using a 2D light scattering microfluidic cytometric technique is described. Laser light is coupled into a microfluidic cytometer via an optical fiber to illuminate a single scatterer in a fluidic flow. The 2D light scattering patterns are obtained by using a charge-coupled device (CCD) detector. The system is tested by using standard polystyrene beads of 4 μm and 9.6 μm in diameter, and the bead experimental results agree well with 1D Mie theory simulation results. Experiments on yeast cells are performed using the microfluidic cytometer. Cell results are studied by finite-difference time-domain (FDTD) method, which can simulate light scattering from non-homogeneous cells. For example, a complex biological cell model with inner mitochondrial distribution is studied by FDTD in this paper. Considering the yeast cell size variations, the yeast cell 2D scatter patterns agree well with the FDTD 2D simulation patterns. The system is capable of obtaining 2D side scatter patterns from a single biological cell which may contain rich information on the biological cell inner structures. The integration of light scattering, microfluidics and fiber optics described here may ultimately allow the development of a lab-on-chip cytometer for label-free detection of diseases at a single cell level.

A novel gene delivery system using magnetic nanodarts

One aspect of gene therapy is the efficient transfection of a cell population with DNAs or RNAs (including siRNAs) in order to modulate the expression of genes, to produce cell types for use in gene therapy, or to meet other very important unmet medical needs. Gene therapy has a huge potential in modifying the blueprints of a human cell and giving additional tools for modern medicine. Gene therapy also offers hope in combating lethal genetic diseases, cancer and certain infectious diseases. According to the report “Meeting the challenge: U.S. Industry faces the 21st century: The U.S. biotechnology industry”, the office of technology policy, U.S. Department of Commerce predicted the growth of this industrial sector to be an estimated

γ-catenin contributes to leukemogenesis induced by AML-associated translocation products by increasing the self-renewal of very primitive progenitor cells

5 Billion within the next decade.