Abstract

The hypotheses that form the basis of this proposal are that protease nexin-2/amyloid beta-protein precursor (PN-2/APP) has a critical role in the physiologic regulation of hemostatic factors XIa and IXa. Human platelets, the major intravascular repository for PN-2/APP, function as the physiologic, intravascular delivery system for this protein. Human brain, enriched with PN-2/APP, utilizes this protein as an important anticoagulant. Abnormal processing an/or expression of PN-2/APP by platelets and cerebral blood vessels contribute to the pathogenesis of Alzheimer's Disease (AD) and Hereditary Cerebral Hemorrhage with Amyloidosis-Dutch Type (HCHWA-D). The objectives of this proposal are to study the effect of PN-2/APP on hemostatic proteins and its expression/processing by human platelets and in cerebral blood vessel tissue.
The specific aims are as follows: (1) Studies will be performed to ascertain the interaction of PN-2/APP with factors Ixa and Xia. The kinetics of factor Ixa inhibition by PN-2/APP will be characterized. Pn- 2/APP inactivation rates of factors Xia and Ixa will be determined in the absence or presence of heparin and platelets or cultured endothelial cells. Investigations will determine if, when in excess, factor Ixa and factor Xia proteolyze PN-2/APP. (2) Investigations will be performed to study the processing and expression of human platelet PN-2/APP. Studies will be performed to determine if upon lysis of human platelets, endogenous calpain produces an amyloidogenic fragment in normal and AD platelets. Investigations will determine if thrombin or platelet activating factor can induce calpain processing of PN-2/APP in the absence of cell lysis. Studies will be conducted to characterize the structure of membrane APP remaining with platelets after PN-2/APP secretion. PN-2/APP will be quantitated in whole lysates and releasates from normal and AD platelets. (3) Investigations will determine the expression and processing of PN-2/APP in cerebral blood vessels. Immunohistochemical and in situ hybridization studies on human brain tissue from HCHWA-D patients, AD patients, and normals will determine the synthesis and form of PN-2/APP in cerebral vessel walls. Pn-2/APP inactivation rates of factors Xia and Ixa in the presence of cultured smooth muscle cells will be determined. Cultured smooth muscle cells will be studied to determine how they express and process PN-2/APP. Better understanding of the biochemistry and physiology of PN-2/APP will provide insight into the patho-biochemistry and patho- physiology of AD and HCHWA-D, two disorders intimately involved with PN- 2/APP.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL049566-03
Application #
2225649
Study Section
Special Emphasis Panel (ZHL1-CSR-J (01))
Project Start
1992-09-30
Project End
1996-08-31
Budget Start
1994-09-30
Budget End
1995-08-31
Support Year
3
Fiscal Year
1994
Total Cost
Indirect Cost

  Institution

Name
University of California Irvine
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
161202122
City
Irvine
State
CA
Country
United States
Zip Code
92697

  Publications

Jung, Sonia S; Zhang, Weibing; Van Nostrand, William E (2003) Pathogenic A beta induces the expression and activation of matrix metalloproteinase-2 in human cerebrovascular smooth muscle cells. J Neurochem 85:1208-15
Davis, Judianne; Wagner, Matthew R; Zhang, Weibing et al. (2003) Amyloid beta-protein stimulates the expression of urokinase-type plasminogen activator (uPA) and its receptor (uPAR) in human cerebrovascular smooth muscle cells. J Biol Chem 278:19054-61
Van Nostrand, William E; Melchor, Jerry P; Keane, David M et al. (2002) Localization of a fibrillar amyloid beta-protein binding domain on its precursor. J Biol Chem 277:36392-8
Wagner, M R; Keane, D M; Melchor, J P et al. (2000) Fibrillar amyloid beta-protein binds protease nexin-2/amyloid beta-protein precursor: stimulation of its inhibition of coagulation factor XIa. Biochemistry 39:7420-7
Mahdi, F; Rehemtulla, A; Van Nostrand, W E et al. (2000) Protease nexin-2/Amyloid beta-protein precursor regulates factor VIIa and the factor VIIa-tissue factor complex. Thromb Res 99:267-76
Van Nostrand, W E; Melchor, J; Wagner, M et al. (2000) Cerebrovascular smooth muscle cell surface fibrillar A beta. Alteration of the proteolytic environment in the cerebral vessel wall. Ann N Y Acad Sci 903:89-96
Hook, V Y; Sei, C; Yasothornsrikul, S et al. (1999) The kunitz protease inhibitor form of the amyloid precursor protein (KPI/APP) inhibits the proneuropeptide processing enzyme prohormone thiol protease (PTP). Colocalization of KPI/APP and PTP in secretory vesicles. J Biol Chem 274:3165-72
Annich, G; White, T; Damm, D et al. (1999) Recombinant Kunitz protease inhibitory domain of the amyloid beta-protein precursor as an anticoagulant in venovenous extracorporeal circulation in rabbits. Thromb Haemost 82:1474-81
Van Nostrand, W E; Porter, M (1999) Plasmin cleavage of the amyloid beta-protein: alteration of secondary structure and stimulation of tissue plasminogen activator activity. Biochemistry 38:11570-6
Van Nostrand, W E; Melchor, J P; Ruffini, L (1998) Pathologic amyloid beta-protein cell surface fibril assembly on cultured human cerebrovascular smooth muscle cells. J Neurochem 70:216-23

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