Project summary
The Candidate is a young investigator dedicated to developing an academic career focused on understanding
the mechanisms by which ethanol disrupts balance between immune cell survival and death pathways. With a
strong background in liver diseases and related innate immunity, the candidate has developed particular
expertise in the use of in vivo and cell culture models to conduct the proposed studies. The Candidate’s current
work has provided her with the opportunity to develop her own research program and begin her transition to
independence. The Career Development Plan outlines 2-years of mentored training on technical skills and
career development activities designed to promote the successful transition to independence. A 3-year program
of independent scientific and career development after successful recruitment as an Assistant Professor position
is also outlined. The Candidate’s Mentor has a proven track-record of excellent scientific productivity and
successful mentorship and can provide the Candidate with a solid research environment in her lab. Research
plan: Alcohol-associated liver disease (ALD) remains a major socioeconomic burden with high morbidity and
mortality. Activation of innate immunity not only contributes to progression of ALD, but is also critical for resolution
of injury. Appropriate regulation of pro-survival and pro-death pathways is critical to the ability of innate immune
cells to rapidly respond to maintain liver homeostasis in ethanol-induced injury. MLKL-mediated necroptosis plays
a dual role in tissue injury/repair. Whether MLKL-mediated necroptosis of immune cells is beneficial or
detrimental in the context of ethanol is unknown. In preliminary work, Mlkl deficiency in myeloid cells exacerbated
ethanol-induced injury, associated with increased ethanol-induced accumulation of hepatic macrophages and
neutrophils. Importantly, we found that Gao-binge increased necrosis/necroptosis of hepatic F4/80+ cells,
suggesting that MLKL may protect from ethanol-induced injury by promoting macrophage death and resolution
of inflammation. Together, these data led us to hypothesize that myeloid cell-specific MLKL restricts ethanol-
induced injury by regulating hepatic immune cell survival and death. To test this hypothesis, hepatic immune cell
populations and death will be characterized by flow cytometric analysis of isolated NPCs from livers of Mlkl BM
chimeras after Gao-binge. We will then challenge cell-specific knock-outs (LysM, Clec4f and MRP8 CRE crossed
with Mlklfl/fl) to ethanol, to distinguish cell-specific contributions of MLKL to mALD. Further, a phospho-proteomics
paired with targeted mechanistic approaches will be utilized to determine whether ethanol-mediated activation
of MLKL is RIP3-dependent and/or RIP3-independent. Additionally, since neutrophils and neutrophil extracellular
traps (NETs) are important in ALD, the project will further explore whether neutrophil-specific MLKL limits
ethanol-induced injury by regulating NETs. Altogether, this proposal will provide a solid foundation for future
mechanistic studies and clinical interventions for ethanol-induced injury and inflammation.