Regulation and function of astrocytic Piezo1 in neuroinflammation and sex-dependent Alzheimer’s disease-like pathologies

Structural tissue alterations in numerous brain disorders can initiate mechanosensory signaling pathways and influence neuropathology. Astrocytes are highly mechanosensitive cells that are essential for maintaining brain homeostasis, yet the mechanisms underlying astrocytic mechanosensation in inflammatory conditions and Alzheimer’s disease (AD) remain poorly understood. In this thesis, I investigated the regulation and functional role of the mechanosensitive ion channel Piezo1 in astrocytes during neuroinflammation and AD pathology. Using primary cell cultures, I found that direct exposure of astrocytes to inflammatory stimuli, including lipopolysaccharide (LPS) or oligomeric amyloid-β (oAβ), had minimal effects on Piezo1 expression. In contrast, when LPS or oAβ were applied to primary microglial cultures, conditioned media from these cultures significantly upregulated Piezo1 expression in astrocytes. I further identified that pro-inflammatory cytokines released by microglia (IL-1α, IL-1β, and TNF-α) can directly enhance Piezo1 expression and Piezo1- mediated Ca²⁺ signaling in both rodent and human astrocytes. In vivo, microglial depletion in 5xFAD mice significantly reduced astrocytic Piezo1 expression. Functional studies showed that Piezo1 activation suppressed inflammatory gene expression under cytokine-primed conditions, whereas Piezo1 knockdown enhanced astrocyte reactivity and promoted inflammatory responses and microglial activation. These findings demonstrate that astrocytic Piezo1 is regulated by microglia-derived inflammatory signals and plays a context-dependent role in modulating astrocyte reactivity and neuroinflammatory responses. To further examine the role of astrocytic Piezo1 in AD, astrocyte-specific Piezo1 conditional knockout (cKO) 5xFAD mice were used. Astrocytic Piezo1 cKO increased glial activation, pro-inflammatory mediator expression, and blood-brain barrier permeability in both sexes. Astrocytic Piezo1 cKO selectively reduced amyloid-β plaque burden in male mice, whereas it increased tau phosphorylation and worsened spatial working memory in female mice, highlighting a sex-specific effect of astrocytic Piezo1 signaling. Bulk RNA sequencing analysis further revealed significant enrichment of inflammatory pathways and negative enrichment of metabolic and energy-related pathways in both sexes following astrocytic Piezo1 cKO. Only female mice showing significant enrichment of apoptosis, complement, and synapse-related pathways. These findings identify astrocytic Piezo1 as a critical regulator of neuroinflammation and reveal sex-dependent effects of astrocytic Piezo1 deletion on AD pathology, suggesting that Piezo1 signaling may be associated with differential disease progression in male and female AD mice.