The miR-183/96/182 cluster is a checkpoint for resident immune cells and shapes the cellular landscape of the cornea

Abstract

PurposeThe conserved miR-183/96/182 cluster (miR-183C) regulates both corneal sensory innervation and corneal resident immune cells (CRICs). This study is to uncover its role in CRICs and in shaping the corneal cellular landscape at a single-cell (sc) level. MethodsCorneas of naïve, young adult [2 and 6 months old (mo)], female miR-183C knockout (KO) mice and wild-type (WT) littermates were harvested and dissociated into single cells. Dead cells were removed using a Dead Cell Removal kit. CD45+ CRICs were enriched by Magnetic Activated Cell Sorting (MACS). scRNA libraries were constructed and sequenced followed by comprehensive bioinformatic analyses. ResultsThe composition of major cell types of the cornea stays relatively stable in WT mice from 2 to 6 mo, however the compositions of subtypes of corneal cells shift with age. Inactivation of miR-183C disrupts the stability of the major cell-type composition and age-related transcriptomic shifts of subtypes of corneal cells. The diversity of CRICs is enhanced with age. Naïve mouse cornea contains previously-unrecognized resident fibrocytes and neutrophils. Resident macrophages (ResMφ) adopt cornea-specific function by expressing abundant extracellular matrix (ECM) and ECM organization-related genes. Naïve cornea is endowed with partially-differentiated proliferative ResMφ and contains microglia-like Mφ. Resident lymphocytes, including innate lymphoid cells (ILCs), NKT and γδT cells, are the major source of innate IL-17a. miR-183C limits the diversity and polarity of ResMφ. ConclusionmiR-183C serves as a checkpoint for CRICs and imposes a global regulation of the cellular landscape of the cornea.