Sickle cell disease (SCD) is the most common genetic hematologic disorder in the United States. The pathophysiology of organ dysfunction in SCD extends beyond abnormal rheology and hemolysis, and includes abnormal vascular function, thrombosis, and inflammation. There is increasing evidence that oxidative stress is an important biochemical trigger for vaso-occlusion (VO) in SCD. Increased reactive oxygen species (ROS) can lead to VO through platelet and leukocyte activation, and microvascular dysfunction. Whether pharmacologic reduction in oxidative stress prevents VO and improves microvascular perfusion is not known. The goal of this proposal is to use state-of-the-art microvascular perfusion and molecular imaging techniques together with advanced plasma biochemical analysis to test mechanism and therapeutic impact of two promising anti-oxidants implicated in reducing ROS in SCD: (a) N-acetylcystine (NAC), and (b) apocynin.
In Aim 1, the Townes murine model of SCD and control mice will be studied. Non-invasive microvascular perfusion imaging with contrast-enhanced ultrasound (CEU) of the kidney and skeletal muscle under normoxic and post-hypoxic conditions will be performed in mice pre-treated for 6 weeks with NAC, apocynin, or vehicle. Parametric perfusion analysis (microvascular blood volume and flux rate) and dispersion modeling will be used to better characterize the vascular events responsible for abnormal flow in the SCD model or treatment effect.
In Aim 2, CEU molecular imaging of activated von Willebrand Factor (vWF) and platelets, and PET molecular imaging of oxidative stress will be used to further assess the microvascular molecular events responsible for abnormal perfusion or treatment effect from the NAC or apocynin. In both of these Aims, advanced LC-MS/MS for unbiased metabolomics and targeted identification of oxidative modification of VWF and its main regulatory protease (ADAMTS-13), as well as other key ROS, will be performed to evaluate drug effect.
In Aim 3, CEU perfusion imaging of limb skeletal muscle, kidney, and myocardium will be performed in a pilot clinical trial testing the hypothesis that reducing oxidative stress with NAC improves microvascular perfusion in patients with SCD. This trial is designed as a double cross-over with NAC or placebo for 6 weeks each. The results of this proposal will address a critical gap in our knowledge of the pathobiology of SCD and provide insight into new therapeutic interventions. It will also provide the PI with a rich research training program (including multidisciplinary mentorship and advisory teams, comprehensive coursework, and career advancement resources available at the institution) that will lay the foundation to become an independent investigator who is able to use advanced imaging and other analytic techniques in pre-clinical and clinical studies to examine microvascular pathophysiology in hematologic disease.
This project will use contrast-enhanced ultrasound blood flow and molecular imaging to study how therapies that target oxidative stress can prevent vaso-occlusion in mice and humans with sickle cell disease. It will also allow the candidate to emerge with a unique set of skills that will lay the foundation to become an independent investigator who can answer important questions in hematologic disease.
|Belcik, J Todd; Davidson, Brian P; Xie, Aris et al. (2017) Augmentation of Muscle Blood Flow by Ultrasound Cavitation Is Mediated by ATP and Purinergic Signaling. Circulation 135:1240-1252|
|Wu, Melinda D; Atkinson, Tamara M; Lindner, Jonathan R (2017) Platelets and von Willebrand factor in atherogenesis. Blood 129:1415-1419|