The objective of this application is to elucidate the antigens that drive the inflammatory response in kidney ischemia reperfusion injury (IRI). Kidney IRI has a significant detrimental effect on native kidneys and allografts. Recent data from our group and others in kidney, liver, lung, intestine and brain has shown a direct role for lymphocytes, particularly CD4 T cells, in IRI. T cells typically respond to alloantigens, or modified self antigens. To date, the antigens that fuel inflammation and tissue injury in IRI are not known. The primary applicant will partner with an experienced protein chemist and basic immunologists to synergize across disciplines for a high risk, high impact, novel line of investigation in kidney IRI. Hypothesis: Self antigens play an important role in driving inflammation and immune responses of T cells in IRI.
Specific Aim 1. We will use an established mouse model of kidney clamp induced warm IRI. We will obtain peri-renal lymph nodes from IRI and sham animals, and use HLA-class II immunoprecipitation, isolate the IRI presented peptides. Using highly sophisticated protein isolation and identification techniques including HPLC/Mass Spectroscopy analysis of MHC-peptide complex, we will identify the specific epitopes of proteins involved in T cell immune responses during renal IRI.
Aim 2. We will inject the synthetic peptide T cell epitopes identified in Aim 1 to naive mice, and at select times, the functional, histologic, molecular and immunological parameters of IRI will be evaluated. We will load isolated synthetic peptides to dendritic cells (DCs) in vitro and then these loaded DCs will be injected into mice. After injecting peptides or loaded DC's the mice will be submitted to IRI to determinate the level of protection and tissue response. Expected results: to find novel peptides that will have therapeutic potential to decrease tissue injury from IRI.
The antigens that drive the inflammatory responses in kidney ischemia reperfusion injury are unknown. We plan to use a combined protein chemistry, immunology and in vivo model approach to identify the peptides in the kidney that are involved in kidney IRI. The study of these can lead to new therapeutics to decrease tissue injury and inflammation after kidney IRI.
