Transplant recipients developing anti-donor HLA antibodies after transplantation demonstrate increased risk of antibody-mediated rejection, transplant associated vasculopathies and graft loss. HLA class I signaling pathways have been implicated in these processes because ligation of class I molecules by anti-HLA antibodies initiates intracellular signals in endothelial cells that synergize with growth factor receptors to elicit survival and proliferation. We found that anti-HLA antibodies elicit different signal transduction outcomes, survival vs. proliferation, depending upon their specificity, titer and the degree of HLA antigen expression on the endothelial cell. These studies provide a paradigm for further elucidating the molecular mechanisms underlying how the concentration of antibody mediates survival and proliferation events in endothelium and their relationship to transplant outcome. Activation of anti-apoptotic and cell survival machinery in endothelial cells is augmented when cells are exposed to low concentrations of anti-HLA antibodies. In contrast, treatment of endothelial cells with high concentrations of HLA antibodies stimulates cell proliferation. This suggests that low levels of antibody binding to HLA may be beneficial to transplant survival by activating survival pathways that promote graft accommodation. On the other hand, high levels of antibody binding may have a detrimental effect on graft survival by upregulating FGFR expression, stimulating cell proliferation and increasing risk for development of transplant vasculopathy. We believe these findings are clinically relevant and may explain differences in transplant outcome in recipients producing anti-donor HLA antibodies. The overall goals of this proposal are to elucidate whether the intracellular signaling events initiated by antibody ligation of class I molecules are influenced by the specificity and concentration of the antibody and to determine the clinical relevance of class I signaling pathways in transplantation.
Aim 1 will focus on HLA interactions with 2 integrins and survival and proliferation phosphorylation cascades induced by different titers of anti-HLA antibody on primary human endothelial cells. These studies will permit us to dissect the class I signaling pathways and explore the cause-effect relationships between proteins in the pathway.
Aim 2 will test whether the antibody induced phosphorylation cascades identified in cultured EC are also operational in vivo. We will study the effect of different titers and specificities of anti-MHC antibody on induction of cell survival and proliferation signal transduction pathways in a RAG1 knock out murine heterotopic cardiac transplant model.
Aim 3 will explore whether anti-MHC class I induced phosphorylation of proteins in cardiac transplant biopsies correlate with the presence and titer of circulating anti-donor-HLA antibodies, diagnosis of antibody-mediated acute and chronic rejection and transplant outcome. These experiments will confirm the biological relevance of class I mediated signaling in human transplantation.

Public Health Relevance

Studying anti-HLA antibody mediated signal transduction in the proposed in vitro and in vivo models will permit us to establish the importance of the class I signaling pathways in solid organ transplant outcome. These studies will identify key signaling proteins mediating acute and chronic antibody-mediated rejection and may permit the development of new treatment strategies. Using samples from transplant recipients, this study will develop and test class I antibody mediated signal transduction as reliable indicators of transplant outcome and provide insight into mechanisms underlying antibody mediated graft injury.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI042819-12
Application #
8207920
Study Section
Atherosclerosis and Inflammation of the Cardiovascular System Study Section (AICS)
Program Officer
Nabavi, Nasrin N
Project Start
1999-01-01
Project End
2014-12-31
Budget Start
2012-01-01
Budget End
2012-12-31
Support Year
12
Fiscal Year
2012
Total Cost
$381,150
Indirect Cost
$133,650
Name
University of California Los Angeles
Department
Pathology
Type
Schools of Medicine
DUNS #
092530369
City
Los Angeles
State
CA
Country
United States
Zip Code
90095
Li, Fang; Wei, Jennifer; Valenzuela, Nicole M et al. (2015) Phosphorylated S6 kinase and S6 ribosomal protein are diagnostic markers of antibody-mediated rejection in heart allografts. J Heart Lung Transplant 34:580-7
Tsai, E W; Reed, E F (2014) MHC class I signaling: new functional perspectives for an old molecule. Tissue Antigens 83:375-81
Valenzuela, Nicole M; McNamara, Jeffrey T; Reed, Elaine F (2014) Antibody-mediated graft injury: complement-dependent and complement-independent mechanisms. Curr Opin Organ Transplant 19:33-40
Roedder, Silke; Sigdel, Tara; Salomonis, Nathan et al. (2014) The kSORT assay to detect renal transplant patients at high risk for acute rejection: results of the multicenter AART study. PLoS Med 11:e1001759
Jin, Y-P; Valenzuela, N M; Ziegler, M E et al. (2014) Everolimus inhibits anti-HLA I antibody-mediated endothelial cell signaling, migration and proliferation more potently than sirolimus. Am J Transplant 14:806-19
Valenzuela, N M; Hong, L; Shen, X-Da et al. (2013) Blockade of p-selectin is sufficient to reduce MHC I antibody-elicited monocyte recruitment in vitro and in vivo. Am J Transplant 13:299-311
Valenzuela, Nicole M; Reed, Elaine F (2013) Antibodies in transplantation: the effects of HLA and non-HLA antibody binding and mechanisms of injury. Methods Mol Biol 1034:41-70
Blumberg, Jeremy M; Gritsch, Hans A; Reed, Elaine F et al. (2013) Kidney paired donation in the presence of donor-specific antibodies. Kidney Int 84:1009-16
Valenzuela, Nicole M; Mulder, Arend; Reed, Elaine F (2013) HLA class I antibodies trigger increased adherence of monocytes to endothelial cells by eliciting an increase in endothelial P-selectin and, depending on subclass, by engaging FcýýRs. J Immunol 190:6635-50
Ziegler, Mary E; Souda, Puneet; Jin, Yi-Ping et al. (2012) Characterization of the endothelial cell cytoskeleton following HLA class I ligation. PLoS One 7:e29472

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