The Human Tissue Acquisition and Processing Core (Scientific Core B) coordinates the acquisition of human tissues from which mast cells, basophils and B cells are obtained. Mast cells derived from tissues and basophils and B cells from peripheral blood, after they have matured in their natural environments, best reflect the functional attributes of these cells in vivo. They lack the chromosomal aberrations and gene expression abnormalities of leukemic cells, and also exhibit key differences from those cells derived in vitro from progenitors. For example, cord blood-derived mast cells express primarily FcyRIIb, an inhibitory receptor, while those dispersed from skin and lung express only FcyRIIa, an activating receptor. Fresh surgical lung and skin will be used as sources of mature human mast cells, peripheral blood for basophils and B cells. The MCT cell is the predominant mast cell type in lung, the MC{TC} cell in skin. The National Diseases Research Interchange (www.ndriresource.org/), Cooperative Human Tissue Network (http://www-chtn.ims.nci.nih.gov/) and Pathology Department at VCD will provide skin and lung;and the Virginia Blood Services (www.vablood.org/) (buffy coat cells) and individual donors will provide a source of peripheral blood. Mast cells will be dispersed from lung and skin, purified with anti-Kit mAb, separated into MCT and MCTc cells with anti-CD88 Ab as needed, placed into culture and distributed to the appropriate Projects. Basophils and B cells from buffy coats or peripheral blood will be purified by density-dependent sedimentation and negative selection, and distributed. Project 1 requires tissue-derived mast cells and peripheral blood basophils to study FceRI and/or FcyRIIa mediated desensitization, and facilitates collaborations with Dr. Spiegel (P4) on the involvement of SphK and CerK and with Dr. Ryan (P3) on the involvement of Lyn in desensitization. Project 2 utilizes B cells from peripheral blood or buffy coats to extend the animal model of CD23-dependent regulation of IgE production to humans by assessing in vitro the effect of novel CD23 modulating agents on IgE production. Project 4 utilizes tissue-derived mast cells to examine the effects of novel inhibitors of sphingosine and ceramide kinases on mediator production by these cells after they are activated, and facilitates collaborations with Drs. Schwartz and Kepley on the roles of these enzymes on human mast cell functions. Core B also will manage the Odyssey Infrared Imaging System, which will be used primarily for processing and analyzing Western blots of the signal transduction molecules under study in Projects 1-4. Thus, Core B relates to each of the proposed Projects.
Showing the most recent 10 out of 56 publications
