A single-cell atlas of normal and KRAS^G12D-malformed lymphatic vessels

Somatic activating mutations in KRAS can cause complex lymphatic anomalies (CLAs). However, the specific processes that drive KRAS-mediated CLAs have yet to be fully elucidated. Here, we used single-cell RNA sequencing to construct an atlas of normal and Kras^G12D-malformed lymphatic vessels. We identified 6 subtypes of lymphatic endothelial cells (LECs) in the lungs of adult wild-type mice (Ptx3, capillary, collecting, valve, mixed, and proliferating). To determine when the LEC subtypes were specified during development, we integrated our data with data from 4 stages of development. We found that proliferating and Ptx3 LECs were prevalent during early lymphatic development and that collecting and valve LECs emerged later in development. Additionally, we discovered that the proportion of Ptx3 LECs decreased as the lymphatic network matured but remained high in Kras^G12D mice. We also observed that the proportion of collecting and valve LECs was lower in Kras^G12D mice than in wild-type mice. Last, we found that immature lymphatic vessels in young mice were more sensitive to the pathologic effects of Kras^G12D than mature lymphatic vessels in older mice. Together, our results expand the current model for the development of the lymphatic system and suggest that KRAS mutations impair the maturation of lymphatic vessels.