An Integrated Neuroimmune Assembloid Model to Advance Neurodegenerative Disease Studies

Abstract Brain organoids are three-dimensional cultures derived from human pluripotent or embryonic stem cells that recapitulate key genetic, biochemical, and molecular features of the human brain. They provide a powerful platform for studying human brain development and modeling genetic neurological disorders. However, their application to age-dependent neurodegenerative diseases remains limited, largely due to the absence of standardized methods for incorporating functional microglia, critical regulators of neuroinflammation and disease progression. Here, we describe a strategy for generating neuroimmune assembloids, brain organoids containing functional glial cells capable of mounting inflammatory responses. By introducing hematopoietic progenitor cells into developing brain organoids, we enable their in situ maturation into microglia-like cells that persist in culture for up to one month. These cells exhibit hallmark microglial behaviors, including morphological remodeling, migration, phagocytosis and transcriptional changes in response to inflammatory stimuli. Together, these immunocompetent-like brain organoids provide a promising and versatile platform for investigating neuroimmune interactions and neuroinflammatory mechanisms underlying age-related neurodegenerative diseases.