(628g) Stiffness Induces Aging-like Phenotypic Changes in Microglia

Aging is associated with tissue regeneration and significant loss of function in brain cells, including microglia. Microglia play a critical role in the primary immune response of the central nervous system which are highly active and motile cells interacting chemically and mechanically with their environment. Studies have shown that brain stiffness is significantly higher in aging brains compared to younger brains. The role of matrix stiffness as related to subtle but pivotal changes in microglia physiology and dysfunction is underexplored. The overall goal of our study is the development and implementation of a platform that enables the convergence of engineered cell microenvironments with the phenotypic and functional analysis of microglia. Using our innovative biomimetic model that allows modulation of substrate stiffness (2 kPa, 8 kPa, 15 kPa, and 25 kPa mimicking young, mid-age, middle-age, and aged brain tissue, respectively), we investigated the role of matrix stiffness in modulating microglial phenotype and function. We demonstrated that stiffness increased microglial proliferation and migration, elevated expression of inflammatory markers, and a heightened inflammatory M1 profile. We also demonstrated that microglia cultured on aging-like stiffness showed 1) increased ROS production, 2) impaired mitochondrial respiration, and 3) increased lipid droplet accumulation and total intracellular cholesterol. We observed a significant increase in oxidized glutathione (GSSG) in microglia cultured on aging-like stiffness compared to young brain stiffness. A similar effect has been observed in microglia isolated from aging murine models indicating a correlation to physiological conditions. Pretreatment with n-acetyl-cysteine (NAC) ameliorates stiffness-induced microglial growth and ROS generation. These data suggest a plausible mechanism that increased stiffness modulates microglial dysfunction. Understanding the impact of stiffness on microglial biology will provide significantly more nuanced data to intervene in an aging-related loss in brain function.