ELUCIDATING THE ROLE OF CRB2 IN VENTRICULAR SYSTEM DEVELOPMENT AND HYDROCEPHALUS PATHOGENESIS

Hydrocephalus involves cerebrospinal fluid accumulation and ventricular enlargement and can result from mutations in CRB2 (crumbs cell polarity complex component 2), which are associated with congenital hydrocephalus, ventriculomegaly, periventricular heterotopias, and aqueduct stenosis in humans. CRB2 is an apical polarity protein implicated in epithelial organization, ciliogenesis, and adherens junction stability, yet how its dysfunction drives hydrocephalus remains unclear, particularly in neural progenitors that generate the ependymal lining. We hypothesized that CRB2 is potentially required in neural progenitors for proper ependymal cell development and ventricular integrity. Using multiple Cre driver mouse lines to conditionally delete Crb2, we found that Crb2 deletion in broad neural progenitors (Nestin-Cre) led to a marked reduction in ependymal and ciliated ventricular lining cells, supporting an early requirement for CRB2 in ependymal generation. In contrast, Crb2 deletion after lineage specification (Foxj1-Cre) produced only mild abnormalities, suggesting a more limited role in ependymal maintenance. Targeting cortical progenitors (Emx1-Cre) revealed disrupted adherens junctions and decreased apical polarity complex proteins at the ventricular surface, consistent with impaired ventricular wall integrity. Because Nestin-Cre mutants develop aqueduct stenosis, we also examined midbrain-specific loss (Wnt1-Cre2) and observed reduced rostral aqueduct ependymal cells, abnormal cilia, and fewer subcommissural organ cells, implicating CRB2 in aqueduct formation. Together, these findings identify CRB2 as essential in neural progenitors for ependymal development and ventricular surface integrity, providing a mechanistic framework for CRB2-linked hydrocephalus.