An official website of the United States government.

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you’ve safely connected to the .gov website. Share sensitive information only on official, secure websites.

COMPLEMENT AND EFFEROCYTOSIS IN CLEARING PYROPTOTIC CELLS

Investigators
Miao, Edward A; Song, Wenchao
Institutions
University of North Carolina
Start date
2018
End date
2023
Objective
ABSTRACT Many pathogens invade host cells and replicate in the protected intracellular niche. The most direct way tocounteract this virulence strategy is to kill the afflicted cell by programmed cell death. Pyroptosis is a form ofprogrammed cell death initiated by caspase-1- or -11-driven opening of the gasdermin pore. Although the hostcell is killed, we have shown both in vivo and in vitro the intracellular bacteria survive the process of pyroptosis.However, we found that instead of being dispersed into the intracellular space, bacteria within pyroptotic cellsbecome trapped in the torn but mostly intact plasma membrane. Because this trapping is immunologicallyuseful to prevent dissemination, and because the structure of a pyroptotic cell is different than the term debris,we chose to name this structure as a ?pore-induced intracellular trap? or PIT. The PIT serves as a nidus for complement deposition, which attracts neutrophils to the PIT. Theneutrophils then efferocytose (phagocytosis of a dead cell) the PIT and the bacteria trapped within. Ultimately itis the neutrophil, therefore, that kills the intracellular bacteria. Salmonella Typhimurium has intestinal virulence factors and intracellular virulence factors. In the intestine,it expresses flagellin and invades intestinal epithelial cells using the SPI1 T3SS. Flagellin and SPI1 are readilydetected by the NLRC4 inflammasome that activates caspase-1. However, during intracellular replication inmacrophages that dominates systemic disease, the bacteria repress flagellin and express the NLRC4-evasiveSPI2 T3SS. In order to study pyroptosis in vivo, we engineered the bacteria to express flagellin on demand. In Aim 1 we continue to use this flagellin engineered S. Typhimurium to study PIT clearance mechanismsin vitro and in vivo. Complement is required for clearance of the PIT and its trapped bacteria in vivo. Wehypothesize that the reason that complement activates on dead cells is in anticipation that they may retaintrapped intracellular bacteria. We investigate the complement initiation pathways that are triggered by the PIT,the importance of C5a and C3a, and the importance of complement opsonization to drive efferocytosis. In Aim 2 we return to wild type S. Typhimurium, asking if the trapping concepts apply to the gastrointestinalphase of infection where these bacteria are detected by NLRC4. We hypothesize that intestinal epithelial cellsthat exfoliate in response to bacterial invasion also form PITs that trap the bacteria. We further hypothesizethat this trapping in the gut lumen is important because it allows infiltrating neutrophils to preferentially targetinvasive bacteria instead of commensal luminal bacteria. We hypothesize that this is accomplished becauseinvasive bacteria are trapped within the exfoliated intestinal epithelial cells.
Funding Source
Nat'l. Inst. of Allergy and Infectious Diseases
Project source
View this project
Project number
1R01AI136920-01A1
Categories
Bacterial Pathogens