Immortalized Human Dermal Microvascular Endothelial Cells

Gαq-R183Q Impairs Endothelial Cell Migration and Angiogenic Sprouting

Capillary malformations (CMs) are caused by somatic mutations in GNAQ (p.R183Q) within endothelial cells; however, the downstream mediators have yet to be determined. Xu's team mapped the Gαq-R183Q phosphoproteome to identify pathogenic pathways and therapeutic targets.

To investigate Gαq-R183Q signaling in endothelial cells, they generated Gαq knockouts (KO) in immortalized human dermal microvascular endothelial cells (HDMECs) and rescued them with lentiviral expression of mTurquoise2-tagged Gαq-WT or Gαq-R183Q (Fig. 1A). Western blot confirmed similar Gαq expression in both rescued lines (~3-fold higher than parental HDMECs) (Fig. 1A, B). Immunofluorescence of VE-cadherin and F-actin showed that Gαq-R183Q induced cell elongation without affecting cell-cell junctions or cytoskeletal organization compared to Gαq-WT (Fig. 1C). Morphometric analysis confirmed significantly larger cell sizes in Gαq-R183Q HDMECs (Fig. 1D). Given that dysregulated endothelial migration and angiogenic sprouting drive vascular malformations, we investigated how Gαq-R183Q affects these functions. Scratch wound assays revealed that Gαq-R183Q significantly impaired HDMEC migration, resulting in delayed wound closure compared to Gαq-WT (Fig. 1E, F). In VEGF-induced spheroid-based sprouting assays, Gαq-R183Q strongly decreased sprout formation and elongation (Fig. 1G, H), confirming that proper Gαq signaling controls endothelial angiogenic behavior.