eNOS Regulates Lymphatic Valve Specification by Controlling β-Catenin Signaling During Embryogenesis in Mice

Area of Science:

• Vascular biology
• Developmental biology
• Cell signaling

Background:

• Lymphatic valves are essential for unidirectional lymph transport, and their dysfunction causes diseases like lymphedema.
• Lymphatic valve development is initiated by mechanotransduction signaling in response to oscillatory lymph flow during embryogenesis.
• Endothelial NO synthase (eNOS) is a known shear stress effector in blood vessels, but its role in lymphatic development is unknown.

Purpose of the Study:

• To investigate the role of eNOS in lymphatic valve development.
• To elucidate the signaling pathways involved in eNOS-mediated lymphatic valve formation.

Main Methods:

• Utilized global Nos3 knockout mice and human dermal lymphatic endothelial cells.
• Examined beta-catenin activation, nuclear translocation, and target protein expression.
• Performed co-immunoprecipitation and proximity ligation assays to assess protein interactions.
• Investigated the effect of Foxo1 gene ablation on lymphatic valve specification.

Main Results:

• Loss of eNOS reduced lymphatic valve specification by 45% and inhibited beta-catenin activation and nuclear translocation.
• Genetic knockout/knockdown of eNOS downregulated beta-catenin target proteins in vivo and in vitro.
• eNOS directly binds beta-catenin, with binding enhanced by oscillatory shear stress.
• Foxo1 gene ablation partially rescued valve specification defects in eNOS knockout mice.

Conclusions:

• Demonstrated a novel, NO-independent role for eNOS in lymphatic valve specification.
• Proposed a mechanism where eNOS directly binds beta-catenin to regulate its nuclear translocation and transcriptional activity.
• Highlighted the importance of eNOS-beta-catenin interaction in lymphatic development.