Vitamin D binding protein (DBP) is an abundant serum protein belonging to the multigene family that includes albumin, alpha- fetoprotein, and alpha-albumin/afamin. Multiple functions have been attributed to DBP. It binds vitamin D sterols facilitating their serum transport; it sequesters G-actin released to the serum after cellular damage and along with plasma gelsolin constitutes the hypothesized """"""""plasma actin-scavenger"""""""" non-immune host defense system; and it activates macrophages and participates in leukocyte chemotaxis in in vitro models. We recently generated a DBP-/- (null) mouse model and characterized its vitamin D metabolism. This mouse line will constitute the model system to determine DBP's in vivo roles in immune function.
Aim 1 will addresses the hypothesis the DBP functions as a macrophage-activating factor in vivo. Macrophages from the peritoneal, pulmonary, bone marrow, and bone (osteoclasts) compartments of DBP-/- and DBP+/+ mice will be compared and characterized for biological functions that define the activated state. The functions attributable to DBP will be carefully dissected from those attributable to its vitamin D sterol delivery function because 1,25(OH)2D is itself a potent macrophage differentiating factor. This will be done by analyzing the actions of a DBP transgene with a discrete mutation in its vitamin D binding domain.
In Aim 2, DBP's role in host defense will be studied using an endotoxin shock model and by studying the response of DBP-/- mice to infections with a set of macrophage intracellular pathogens.
In Aim 3, the DBP chemotaxis domain will be mapped functionally by analysis of site-directed mutations in vivo, DBP's ability to signal through Ca2+ mobilization will be explored, and the role of megalin in macrophage uptake of DBP will be determined.
In Aim 4, the actin-scavenger hypothesis will be tested by quantifying pulmonary damage in DBP-/-, GSN-/-, and DBP-/-GSN-/- mice after iv G-actin overload. These studies, utilizing a combination of in vitro, cell-based, and in vivo models, will document novel, non-redundant roles for DBP in inflammation and in immune as well as non-immune host defense.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM032035-19
Application #
6604110
Study Section
Orthopedics and Musculoskeletal Study Section (ORTH)
Program Officer
Marino, Pamela
Project Start
1984-04-01
Project End
2005-06-30
Budget Start
2003-07-01
Budget End
2005-06-30
Support Year
19
Fiscal Year
2003
Total Cost
$313,830
Indirect Cost
Name
University of Pennsylvania
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
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