RNA-seq analysis of non-vascular smooth muscle cells (NVSMCs) isolated from the lung, intestine and bladder

To address this, we focused on the Chrm2 gene, which encodes the M2 muscarinic acetylcholine receptor (M2R), a G protein-coupled receptor (GPCR) previously reported to be expressed selectively in NVSMCs and also in CD45-positive immune cells. In contrast, Acta2 is broadly and strongly expressed in all smooth muscle cells (SMCs), including both vascular and non-vascular populations.

To achieve specific labeling of NVSMCs, we generated NVSMC-effector mice by combining Chrm2-Dre with Acta2-Rox-CreER and R26-LoxP-GFP alleles. Following tamoxifen administration, we observed robust GFP fluorescence specifically in NVSMCs of the lung, stomach, and intestine. Notably, GFP labeling was virtually absent in vascular SMCs across multiple organs, confirming the specificity of this genetic strategy.