I propose that during sepsis decreased peritubular capillary perfusion leads to a pro-oxidant peritubular microenvironment, which favors the generation of reactive oxygen species (ROS) and reactive nitrogen species (RNS). My preliminary data show that acute treatment with the naturally occurring anti-oxidant polyphenol, resveratrol, prevented the generation of tubular RNS and the decline in peritubular capillary perfusion observed at 18h following induction of sepsis using the cecal ligation and puncture (CLP) in a murine model of sepsis. Moreover, I have generated direct evidence that resveratrol can scavenge RNS. Based on these findings, I hypothesize that the RNS scavenging ability of resveratrol will protect the mouse renal tubules from the pro-oxidant peritubular microenvironment produced during CLP-induced.
Aim 1 will determine the efficacy of resveratrol to protect renal tubular epithelia from RNS using primary cultures of mouse renal tubular epithelial cells.
Aim 2 will examine the therapeutic potential of resveratrol to prevent tubular epithelial cell injury caused by CLP. Dose-response curves for resveratrol over the time course leading to renal injury will be generated and analyzed for the effects on serum cytokine profile, functional peritubular capillary density, volumetric flow, tubular oxidant generation (each quantified simultaneously using intravital videomicroscopy), and renal injury (morphology and serum BUN/creatinine).
Aim 3 will establish RNS as the key mediator of CLP-induced renal injury in the mouse using iNOS mice and in mice that over-express superoxide dismutase (S0D1) to reduce the levels of NO and O2"""""""" available for RNS (ONOO"""""""") generation, respectively. A very innovative aspect of this study is the use of intravital video microscopy (IWM) to study in real time the mechanisms by which capillary dysfunction develops and leads to tubular injury following CLP. This research proposal will contribute significant to my overall training to become a translational clinical scientist. Each year and more than 210,000 people die from sepsis-related complications in the United States. Acute kidney injury occurs in 20%-50% of septic patients and the mortality of these septic patients is extremely high (~75%) because current therapy is mostly supportive and largely ineffective. The major obstacle to the development of new therapeutic approaches for sepsis-induced kidney injury is the lack of knowledge regarding critical events that can be targeted after the initiation of sepsis. This proposal investigates new therapeutic targets to treat sepsis and has the potential to identify new agents to reduce mortality of this devastating condition.
|Holthoff, Joseph H; Wang, Zhen; Patil, Naeem K et al. (2013) Rolipram improves renal perfusion and function during sepsis in the mouse. J Pharmacol Exp Ther 347:357-64|
|Holthoff, Joseph H; Wang, Zhen; Seely, Kathryn A et al. (2012) Resveratrol improves renal microcirculation, protects the tubular epithelium, and prolongs survival in a mouse model of sepsis-induced acute kidney injury. Kidney Int 81:370-8|
|Wang, Zhen; Holthoff, Joseph H; Seely, Kathryn A et al. (2012) Development of oxidative stress in the peritubular capillary microenvironment mediates sepsis-induced renal microcirculatory failure and acute kidney injury. Am J Pathol 180:505-16|
|Seely, Kathryn A; Holthoff, Joseph H; Burns, Samuel T et al. (2011) Hemodynamic changes in the kidney in a pediatric rat model of sepsis-induced acute kidney injury. Am J Physiol Renal Physiol 301:F209-17|
|Holthoff, Joseph H; Woodling, Kellie A; Doerge, Daniel R et al. (2010) Resveratrol, a dietary polyphenolic phytoalexin, is a functional scavenger of peroxynitrite. Biochem Pharmacol 80:1260-5|