The impact of obesity on neutrophil swarming in septic surgical patients

Project Abstract While obesity has been shown to increase mortality in sepsis, the mechanism underlying this phenomenon is unclear. Dr. Jalilvand’s preliminary data demonstrates that obesity is associated with increased 90-day mortality in surgical patients with sepsis. Further, we have shown that obese adipocytes produce distinct adipocyte-

derived extracellular vesicles (AdEVs) that activate neutrophils and induce disordered neutrophil swarming. These AdEVs are enriched with miR-223, which interacts with PIK3C2A to downregulate the phosphoinositide- 3 kinase (PI3K) pathway, thus reudcing neutrophil migration and chemotaxis. The governing hypothesis for this

proposal is that AdEV-mediated delivery of miR-223 to peripheral neutrophils in obesity impairs neutrophil swarming by modulating the PI3K pathway, thereby contributing to disrupted pathogen clearance in surgical patients with sepsis. We will test this hypothesis with the following aims. AIM 1: Define the role of AdEV-mediated

delivery of miR-223 on PI3K activation and swarming in healthy neutrophils and the efficacy of targeting this pathway to restore swarming in septic obese and lean patients in vitro. We will determine the in vitro differential impact of modulating AdEV miR-223 content on neutrophil function and whether pharmacologic upregulation of

the PI3K pathway can restore swarming in neutrophils isolated from healthy and septic obese and lean individuals. AIM 2: Establish the efficacy of inhibiting miR-223 mediated PI3K downregulation in neutrophils to improve swarming and survival in obese mice with intra-abdominal sepsis. Using a murine model of diet-induced

obesity (DIO) and intra-abdominal sepsis (cecal ligation puncture, CLP), we will test whether systemic delivery of AdEVs enriched with anti-miR-223 improves neutrophil function, bacterial clearance, and survival following CLP in vivo. Lastly, using a conditional, neutrophil-specific knock out of PIK3C2A, we will establish whether

transient silencing of PIK3C2A mitigates miR-223-mediated downregulation of neutrophil PI3K activation and swarming in DIO mice after CLP. While capitalizing on her PhD in adipose tissue inflammation and obesity, this proposal represents a departure from both Dr. Jalilvand and her primary mentor’s, Dr. Hsueh, prior research

focus. A multidisciplinary approach has been crafted to study the proposed aims to develop Dr. Jalilvand’s expertise in EV biology and cargo, live cell imaging to follow neutrophil behavior and quantitative function, pharmacologic targeting, and murine models of sepsis. Completion of this proposal will provide crucial technique

expansion needed to establish a unique area of research distinct from Dr. Jalilvand’s PhD, accomplishing the short-term goals of this career development plan. Paired with the institutional commitment to her success that includes laboratory space, protected time, and laboratory funding, a well-established committee of mentors has

been selected to ensure that Dr. Jalilvand will achieve the long-term goal of becoming an independent surgeon scientist evaluating the implications of obesity in surgical patients with sepsis. As such, this proposal will provide Dr. Jalilvand with the requisite mentorship and resources to transition to independence.