Date on Master's Thesis/Doctoral Dissertation
12-2024
Document Type
Doctoral Dissertation
Degree Name
Ph. D.
Department
Physiology and Biophysics
Degree Program
Physiology and Biophysics, PhD
Committee Chair
Smith, Jason W.,
Committee Member
Metz, Cynthia
Committee Member
Klein, Jon
Committee Member
Maldonado, Claudio
Committee Member
Joshua, Irving
Author's Keywords
shock; glomerulus; proteomics; resuscitation; plasma
Abstract
Introduction: Acute renal dysfunction following traumatic injury and hemorrhagic shock carries significant morbidity and mortality. Despite an extensive understanding of renal tubule damage after injury, less is known regarding early alterations to the glomerular filtration barrier following hemorrhagic shock and resuscitation. Previously, our lab and others have demonstrated improved hepatic and intestinal microcirculatory parameters when resuscitation strategies incorporate fresh frozen plasma. In addition to improving end-organ perfusion, plasma reconstitutes the endothelial glycocalyx, an essential regulator of vascular permeability and microcirculatory function. However, the impact of plasma resuscitation on the glomerular glycocalyx and barrier following hemorrhage remains unknown. We hypothesized that hemorrhagic shock would induce drastic alterations in renal hemodynamics and inflammatory mediator burden resulting in glomerular damage and dysfunction. Furthermore, we hypothesized that resuscitation with plasma (compared to conventional isotonic fluids) would reduce inflammatory mediator burden, replenish the glomerular glycocalyx, and attenuate hemorrhage-induced glomerular barrier damage and dysfunction. Methods: Male, Sprague-Dawley rats (250-300 grams) were subjected to pressured controlled hemorrhagic shock at 40% of baseline mean arterial pressure for 60 minutes, followed by resuscitation with shed whole blood and either lactated ringers (LR) or fresh frozen plasma (FFP). Experimental groups for comparison included: a) Baseline, b) Hemorrhagic shock alone, c) Lactated ringer’s resuscitation, and d) Fresh frozen plasma resuscitation. Renal blood flow and diameter were monitored using doppler ultrasonography. Renal cortex inflammation was evaluated via cytokine and chemokine profiling and histologic identification of early neutrophil and macrophage infiltration. Urine protein concentration was measured as a surrogate for glomerular function and the urine proteome was evaluated to identify markers of glomerular barrier damage and dysfunction following hemorrhagic shock and resuscitation. Selected markers of glomerular damage were confirmed via enzyme-linked immunosorbent assays (ELISA) and glomerular immunohistochemical staining. To assess for renal tubule contributions to alterations in protein handling, damage markers were evaluated in the urine proteome and key proteins involved in protein reabsorption within the proximal convoluted tubule (megalin, cubilin) were measured. Results: Hemorrhagic shock induced a reduction in maximum kidney diameter and lowered renal artery and parenchymal blood flow. Plasma resuscitation resulted in elevated inflammatory cytokines and chemokines in the renal cortex and promoted increased migration of neutrophils and macrophages when compared to resuscitation with LR. Elevated urine protein concentration was seen following hemorrhage and worsened despite resuscitation and restoration of central hemodynamics with LR. However, FFP restored urine protein levels back to baseline. Urinary proteomics identified markers of glomerular damage from all layers of the filter apparatus (ex. hyaluronidase-1, podocalyxin, vinculin). Specifically, plasma resuscitation restored podocalyxin levels back to baseline and attenuated hemorrhaged induced shedding of hyaluronic acid from the glomerular glycocalyx. Urinary markers of tubule damage were not elevated following hemorrhagic shock and resuscitation. Likewise, the concentration of proteins responsible for the reabsorption of protein from the ultrafiltrate did not vary between experimental groups. Conclusions: Hemorrhagic shock and resuscitation results in glomerular dysfunction through damage to multiple components of the glomerular barrier. The resulting proteinuria and alterations of the urine proteome occur in the setting of normal proximal convoluted tubule protein handling. Plasma resuscitation attenuates hemorrhage induced glomerular damage and dysfunction via augmentation of hyaluronic acid and podocalyxin.
Recommended Citation
Risinger, William Byron, "Plasma resuscitation attenuates hemorrhage-induced glomerular barrier damage and dysfunction." (2024). Electronic Theses and Dissertations. Paper 4492.
Retrieved from https://ir.library.louisville.edu/etd/4492
