Harry L Malech

Small numbers of CD34+ primitive hematopoietic progenitors are found in normal human peripheral blood. These cells differentiate to myeloid or lymphoid lineage under the influence of different growth factors. We investigated the effects of IL5 and other growth factors on the production of eosinophils from peripheral blood CD34+ cells. CD34+ cells were enriched from normal donors by apheresis and positive selection using an affinity column and plated in agarose with different combinations ...

Cystic fibrosis (CF) airways are characterized by bacterial infections, excess mucus production, and robust neutrophil recruitment. The main CF airway pathogen is Pseudomonas aeruginosa. Neutrophils are not capable of clearing the infection. Neutrophil primary granule components, myeloperoxidase (MPO) and human neutrophil elastase (HNE), are inflammatory markers in CF airways, and their increased levels are associated with poor lung function. Identifying the mechanism of MPO and HNE ...

Orientation of nucleus, centriole, microtubules, and microfilaments within human neutrophils in a gradient of chemoattractant (5 percent Escherichia coli endotoxin-activated serum) was evaluated by electron microscopy. Purified neutropils (hypaque-Ficoll) were placed in the upper compartment of chemotactic chambers. Use of small pore (0.45 mum) micropore filters permitted pseudopod penetration, but impeded migration. Under conditions of chemotaxis with activated serum beneath the filter, ...

In previous studies amphotropic MFGS-gp91phox (murine onco-retrovirus vector) was used in a clinical trial of X-linked chronic granulomatous disease (X-CGD) gene therapy to achieve transient correction of oxidase activity in 0.1% of neutrophils. We later showed that transduced CD34+ peripheral blood stem cells (CD34+ PBSCs) from this trial transplanted into nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice resulted in correction of only 2.5% of human neutrophils. However, ...

Chronic granulomatous disease (CGD) is a rare recessive disorder caused by defects in the NADPH oxidase enzyme complex of phagocytes (neutrophils, eosinophils and monocytes). CGD phagocytes fail to produce superoxide and other reactive oxygen species following cell activation (Malech, 1993). The products of oxidase activation can be measured in individual cells by flow cytometry using specific fluorescent probes that increase fluorescence upon oxidation (Trinkle et al., 1987). This ...

Reactive oxygen species (ROS) serve several physiological functions; in some settings they act in host defense, while in others they function in cellular signaling or in biosynthetic reactions. We studied the expression and function of a recently described source of ROS, NAD(P)H oxidase 1 or Nox1, which has been associated with cell proliferation. In situ hybridization in mouse colon revealed high Nox1 expression within the lower two-thirds of colon crypts, where epithelial cells undergo ...

BACKGROUND: The treatment of chronic granulomatous disease with conventional allogeneic hematopoietic stem-cell transplantation carries a high risk of serious complications and death. We investigated the feasibility of stem-cell transplantation without ablation of the recipient's bone marrow.

METHODS: Ten patients, five children and five adults, with chronic granulomatous disease underwent peripheral-blood stem-cell transplantation from an HLA-identical sibling. We used a ...

X-linked chronic granulomatous disease (X-CGD) is a primary immunodeficiency caused by mutations in the CYBB gene encoding the phagocyte nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase catalytic subunit gp91(phox). A recent clinical trial for X-CGD using a spleen focus-forming virus (SFFV)-based γ-retroviral vector has demonstrated clear therapeutic benefits in several patients although complicated by enhancer-mediated mutagenesis and diminution of effectiveness over time due ...

Chronic granulomatous disease, a syndrome of recurrent infections and failure of oxidative microbicidal activity in phagocytes, results from defects in the gene for one of several components of an oxidase system that can undergo activation. To determine the relative prevalence of certain of the genetic variants of this disorder, we used immunoblotting to detect two specific neutrophil cytosolic proteins of 47 and 67 kd recently shown to be required for oxidase activation. Chronic ...

BACKGROUND: Previous investigations provide evidence that an enzyme related to the phagocyte NADPH oxidase produces superoxide in the blood vessel wall. These data, however, are confounded by observations that both NADPH and NADH serve as substrates for superoxide production in vascular cells. To clarify this issue, we compared the superoxide-generating capabilities of vascular smooth muscle cells (VSMCs) derived from wild-type (p47phox(+/+); phagocyte oxidase) mice with those from mice ...

OBJECTIVE: Internal tandem duplication (ITD) mutations of the FLT3 receptor are associated with a high incidence of relapse in acute myeloid leukemia (AML). Expression of the CXCR4 receptor in FLT3-ITD-positive AML is correlated with poor outcome, and inhibition of CXCR4 was shown to sensitize AML blasts toward chemotherapy. The aim of this study was to evaluate the impact of FLT3-ITD on cell proliferation and CXCR4-dependent migration in human hematopoietic progenitor cells and to ...

BACKGROUND: The hyper-IgE syndrome (or Job's syndrome) is a rare disorder of immunity and connective tissue characterized by dermatitis, boils, cyst-forming pneumonias, elevated serum IgE levels, retained primary dentition, and bone abnormalities. Inheritance is autosomal dominant; sporadic cases are also found.

METHODS: We collected longitudinal clinical data on patients with the hyper-IgE syndrome and their families and assayed the levels of cytokines secreted by stimulated leukocytes ...

HIV-1-derived lentivectors are promising for gene transfer into hematopoietic stem cells but require preclinical in vivo evaluation relevant to specific human diseases. Nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice accept human hematopoietic stem cell grafts, providing a unique opportunity for in vivo evaluation of therapies targeting human hematopoietic diseases. We demonstrate for the first time that hematopoietic stem cells from patients with X-linked chronic ...

We compared the cell cycle status and expression of mRNA for the amphotropic retroviral receptor in hematopoietic stem cells isolated from bone marrow and cytokine mobilized peripheral blood. CD34+ cells from six normal volunteers were enriched by immune selection from steady-state bone marrow and granulocyte colony-stimulating factor (G-CSF) mobilized peripheral blood (10 microg/kg/day for 5 days). Cell cycle status of the phenotypically primitive CD34+CD38- hematopoietic stem cell ...

Neutrophil extracellular traps (NETs) are implicated in autoimmunity, but how they are generated and their roles in sterile inflammation remain unclear. Ribonucleoprotein immune complexes (RNP ICs), inducers of NETosis, require mitochondrial reactive oxygen species (ROS) for maximal NET stimulation. After RNP IC stimulation of neutrophils, mitochondria become hypopolarized and translocate to the cell surface. Extracellular release of oxidized mitochondrial DNA is proinflammatory in ...