Elucidating mechanisms informing age-related changes in neutrophil function and migration

About This Grant

Project Summary/Abstract The US population aged 65+ is expected to increase to 23% by the year 2050. Age-related chronic inflammatory disease and infections are a significant burden on the healthcare system. Age-related changes in neutrophil function and migration play a significant role in these pathologies. A current gap in understanding mechanisms that mediate age-related changes in neutrophils limits therapeutic intervention. We have developed a scale explant assay using zebrafish across the lifespan to investigate changes in neutrophil function and migration with age in situ. Application of optical metabolic imaging (OMI) provides metabolic readouts in tissues in real time with aging. The Huttenlocher lab has pioneered genetically tractable human iPSC-derived neutrophils (iNeutrophils) to dissect signaling mechanisms that regulate neutrophils. Derivation of iNeutrophils from donors of different ages will enable investigation of these mechanisms with aging. This fellowship proposal will leverage the combined use of these techniques to disentangle the contribution of neutrophil intrinsic and environmental contributions to age-related changes in neutrophil function and migration. We hypothesize that aging alters neutrophil metabolism and function through both neutrophil intrinsic and extrinsic factors. Aim 1 will establish the zebrafish scale explant assay as a model to study aging related inflammation in epithelial tissues, canonical senescence markers, and alterations in peripheral immune cell population. Simultaneously, use of scale explant system and iNeutrophils will elucidate neutrophil metabolic changes in situ and in vitro across the lifespan. Building on our prior work showing a role for neutrophil Lyn kinase in directing migration up an H2O2 gradient, aim 2 will elucidate the role of redox-sensitive Lyn kinase in age-related neutrophil aberrant migration in situ and in vitro. We will introduce a redox-insensitive or wild-type (WT) Lyn kinase into a Lyn-deficient zebrafish. Using the scale explant assay, we will interrogate Lyn’s redox-signaling role in age-related neutrophil chemokinesis and chemotaxis in situ. Depletion of LYN and rescue with redox insensitive or WT LYN in the iNeutrophil system will elucidate the role of Lyn redox-signaling in vitro. We will interrogate how extrinsic H2O2 informs iNeutrophil chemokinesis and chemotaxis. The goal of this work is to identify key mechanisms that regulate age-related changes in neutrophil function for potential therapeutic intervention. This proposal will provide specific training in metabolic imaging, stem cell culture, and cell signaling. This project synthesizes the cell biology expertise of Dr. Huttenlocher with the metabolic and engineering expertise of Dr. Skala along with the vast scientific, medical, and translational resources available at the University of Wisconsin-Madison. This fellowship will promote the development of research, clinical, translational, mentorship, and communication skills crucial for my career as an independent physician scientist.