MODULATION OF PHAGOCYTE ACTIVITY BY MONOCLONAL ANTIBODIE

Project Details

Description

Monoclonal antibodies that interact with functional antigens on
phagocytes are particularly useful as specific probes to study the
role of surface structures in cell activation. By fusing spleen
cells from mice immunized with human peripheral blood
granulocytes with mouse myeloma cells, we have developed a
highly specific monoclonal antibody, DL1.2, that induces
chemotaxis in neutrophilic granulocytes (PMN). DL1.2 also
stimulates f-met-leu-phe (fMLP)-induced chemotaxis. By means
of flow cytometry and immunofluorescence, DL1.2 was found to
react predominantly (greater than 85%) with mature PMN and not
myeloid or monocytic precursor cells. DL1.2 binds to a 120 kD
protein on PMN that we have purified by affinity chromatography.
The overall objective of this proposal is to further characterize
the chemotactic antigen on PMN that binds DL1.2 and to
elucidate the mechanism by which it stimulates chemotaxis. Our
hypothesis is that DL1.2 directly activates a tyrosine specific
protein kinase resulting in phosphorylation of critical membrane
proteins and cytoskeletal elements which subsequently leads to
directed migration. Preliminary studies indicate that DL1.2
induces tyrosine specific protein phosphorylation in PMN, but has
no effect on mobilization of calcium from intracellular stores.
Furthermore, the effects of DL1.2 on PMN are not inhibited by
the guanine nucleotide binding protein inhibitor, pertussis toxin.
Thus, DL1.2, like phorbol myristate acetate, may bypass the
calcium requiring segment of the activation sequence in PMN by
acting directly on a protein kinase. To test this hypothesis, we
plan to examine the effects of DL1.2 alone and in combination
with fMLP on tyrosine kinase activity and protein phosphorylation
in PMN. We will also study the effects of the antibody on
intracellular calcium mobilization and on phosphatidylinositol
turnover since these processes have been linked to
chemoattractant induced PMN activation. Studies on DL1.2 and
its relationship to other antibody binding sites may help to
elucidate the transduction signals that regulate PMN migration.
StatusFinished
Effective start/end date7/1/836/30/92

Funding

  • National Institutes of Health
  • National Institutes of Health
  • National Institutes of Health
  • National Institutes of Health

ASJC

  • Medicine(all)
  • Immunology and Microbiology(all)

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