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Principal Investigator: Samit Ghosh
Organization: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
Fiscal Year: 2024
Award: $608,926
Funding agency: National Institute of Diabetes and Digestive and Kidney Diseases
PROJECT SUMMARY/ABSTRACT
Chronic kidney disease (CKD), widespread among individuals with sickle cell disease (SCD), is a significant
contributor to morbidity and early mortality. The progressive deterioration of renal health in SCD is associated
with chronic intravascular hemolysis. However, the mechanistic approach to interpreting the hemolysis-driven
pathogenesis of CKD in SCD is limited and therefore targeted therapies based on mechanistic models are
lacking. Progressive loss of renal function, represented by increased albuminuria and reduced glomerular
filtration rate (GFR), is associated with microvascular congestion in humans and mice with SCD. We postulated
that circulating cell-free heme, a product of hemolysis, may instigate persistent endothelial damage that ensues
CKD development. Endothelial protein C receptor (EPCR) maintains endothelial barrier integrity. We discovered
an age-dependent as well as heme-induced loss of EPCR from renal microvascular endothelium in SCD (SS)
mice. Concomitantly, the cleaved soluble form of EPCR (sEPCR) was elevated in the plasma associated with
albuminuria. Using super-resolution ultrasound imaging, we found that younger SS mice challenged with the
repetitive infusion of minimal doses of heme and the older SS mice without heme challenge incur substantial
renal microvascular rarefaction. Heme induces endothelial P-selectin that promotes vascular congestion. Our
pilot data show that reduced EPCR expression is associated with increased P-selectin on renal microvascular
endothelium in the SS mice. Proteinase 3 (PR3), expressed on neutrophils, can degrade EPCR. In our repeated
heme-challenged younger SS mice compared to vehicle-injected mice, we noticed increased expression of PR3
in plasma as well as in accumulated neutrophils from the kidneys. Moreover, we found that heme induces
acetylation of histone 4 Lys 16 residues (H4K16ac) which are known to promote the biosynthesis of PR3. Based
on these preliminary data, we hypothesize that heme-induced P-selectin and PR3 concurrently result in the loss
of EPCR leading to endothelial disintegration, microvascular congestion, and CKD development in SCD. We will
test this hypothesis with three specific aims that integrate experiments with human renal endothelial cells and
neutrophils in vitro, murine models, and clinical samples from multiple cohorts of SCD patients.
Aim 1 will determine that repeated low-dose heme insults promote renal endothelial P-selectin that suppresses
EPCR leading to vascular damage and development of CKD in mice with SCD. Aim 2 will test whether heme-
activated PR3 facilitates EPCR cleavage and expedite CKD in SCD. Aim 3 will determine whether sEPCR and
specific genetic variants of EPCR can indicate the risk of CKD in SCD patients.
Terms: <21+ years old><APC receptor><Abnormal Erythrocytes><Abnormal Red Blood Cell><Acceleration><Adhesion Molecule><Adult><Adult Human><Affect><Albuminuria><Anabolism><Anemia><Azurophil Granule Protein 7><Binding><Biological Markers><Blood Neutrophil><Blood Plasma><Blood Polymorphonuclear Neutrophil><Blood Vessels><Bone Marrow Grafting><Bone Marrow Transplant><Bone Marrow Transplantation><C-ANCA><C-ANCA Antigen><CD62P Antigens><Cell Adhesion Molecule Gene><Cell Adhesion Molecules><Cell Body><Cell membrane><Cells><Characteristics><Childhood><Chlorohemin><Chronic><Chronic Kidney Failure><Chronic Renal Disease><Chronic Renal Failure><Clinical><Clinical Data><Cytoplasmic Membrane><DNA><Data><Deoxyribonucleic Acid><Deterioration><Development><Disease Progression><Dose><EPCR><EPCR gene><ESI gene><ESI protein><Echography><Echotomography><Elastase-Specific Inhibitor><Endothelial Cells><Endothelial protein C receptor><Endothelium><Epidemiologic Research><Epidemiologic Studies><Epidemiological Studies><Epidemiology Research><Esteroproteases><Event><Ferriheme Chloride><Ferriprotoporphyrin IX Chloride><Ferroprotoporphyrin><Flow Cytofluorometries><Flow Cytofluorometry><Flow Cytometry><Flow Microfluorimetry><Flow Microfluorometry><Foundations><Frequencies><GMP-140><Gene variant><Genotype><Glomerular Filtration Rate><HDAC Agent><HDAC inhibitor><Hb SS disease><HbSS disease><Health><Heme><Hemin><Hemoglobin S Disease><Hemoglobin sickle cell disease><Hemoglobin sickle cell disorder><Hemolysis><Histone Acetylation><Histone Deacetylase Inhibitor><Histone deacetylase inhibition><Human><Immunoglobulin Enhancer-Binding Protein><Impairment><In Vitro><Individual><Infusion><Infusion procedures><Injections><Intravenous><Kidney><Kidney Urinary System><L-Lysine><LECAM-3><Lead><Leukocyte Proteinase 3><Lysine><Marrow Neutrophil><Marrow Transplantation><Measures><Medical Ultrasound><Mice><Mice Mammals><Microalbuminuria><Modeling><Modern Man><Molecular><Molecular Interaction><Morbidity><Morbidity - disease rate><Murine><Mus><Mutant Strains Mice><Myeloblastin><N-proteinase 4><NF-kB><NF-kappa B><NF-kappaB><NFKB><NP-4><NP4(3)><Neutrophil Proteinase 4><Neutrophil Serine Proteinase><Neutrophilic Granulocyte><Neutrophilic Leukocyte><Nuclear Factor kappa B><Nuclear Transcription Factor NF-kB><Organ><Organ failure><P-Selectin><PI3><PI3 gene><PR-3><Pathogenesis><Patients><Pb element><Peptidases><Peptide Hydrolases><Permeability><Phenocopy><Pilot Projects><Plasma><Plasma Membrane><Plasma Serum><Platelet alpha-Granule Membrane Protein><Polymorphonuclear Cell><Polymorphonuclear Leukocytes><Polymorphonuclear Neutrophils><Predisposition><Progressive Disease><Prolastin><Prophylactic treatment><Prophylaxis><Protease Gene><Proteases><Proteinase 3><Proteinase Inhibitor 3><Proteinases><Proteins><Proteolytic Enzymes><Protoheme><Protohemin><Receptor Gene><Renal Vascular><Renal function><Renal vessels><Research><Reticuloendothelial System, Serum, Plasma><Risk><SKALP><Sampling><Selectins><Severities><Sickle Cell Anemia><Single Base Polymorphism><Single Nucleotide Polymorphism><Specificity><Susceptibility><TRAPPIN 2><Testing><Transcription Factor NF-kB><Transgenic Organisms><Tube><Ultrasonic Imaging><Ultrasonogram><Ultrasonography><Ultrasound Diagnosis><Ultrasound Medical Imaging><Ultrasound Test><Variant><Variation><Vascular Endothelium><Vascular Permeabilities><Wegener Autoantigen><Wegener Granulomatosis Autoantigen><activated protein C receptor><adulthood><age associated><age correlated><age dependent><age linked><age related><age specific><aged><allele variant><allelic variant><bio-markers><biologic marker><biomarker><biomarker discovery><biosynthesis><cell adhesion protein><chronic kidney disease><cohort><developmental><diagnostic ultrasound><disease risk><disorder risk><drug discovery><elafin><endothelial cell protein C receptor><endothelial dysfunction><epidemiologic investigation><epidemiology study><erythrolysis><experiment><experimental research><experimental study><experiments><extracellular vesicles><ferroheme><flow cytophotometry><genetic variant><genomic variant><heavy metal Pb><heavy metal lead><infusions><kappa B Enhancer Binding Protein><kidney function><kidney vascular><kidney vascular structure><mortality><mouse model><mouse mutant><murine model><neutrophil><neutrophil proteinase 3><new marker><novel biomarker><novel marker><nuclear factor kappa beta><overexpress><overexpression><p29><p29b><pediatric><pilot study><plasmalemma><premature><prematurity><prevent><preventing><receptor expression><renal><renovascular><sickle cell disease><sickle cell disorder><sickle disease><sicklemia><sickling><single nucleotide variant><skin-derived antileukoproteinase><sonogram><sonography><sound measurement><super high resolution><superresolution><targeted drug therapy><targeted drug treatments><targeted therapeutic><targeted therapeutic agents><targeted therapy><targeted treatment><therapeutic target><transgenic><ultra high resolution><ultrasound imaging><ultrasound scanning><vascular>