Coordinated lymphangiogenesis is critical in lymph node development and maturation

Background: Lymph node (LN) formation requires multiple but coordinated signaling from intrinsic and extrinsic cellular components during embryogenesis. However, the contribution and role of lymphatic vessels (LVs) in LN formation and maturation are poorly defined. Here, using lymphatic-specific reporters, Prox1-GFP mice and Vegfc(+/LacZ) mice, we analyzed migration, assembly, and ingrowth of lymphatic endothelial cells (LECs) in LNs during pre- and postnatal development. Results: Prox1(+) LECs form string-like connections rather than lymph sac-like structures until E14.5, but the LEC coverage around LN anlagen completes before birth. Compared to wild-type littermates, Vegfc(+/LacZ) mice had markedly smaller LNs in neonates and adults, presumably due to the decrease in LTi cell clusters and migrating Prox1(+) LECs during embryogenesis. In addition, Vegfc-haploinsufficiency or inhibition of VEGFR3 signaling led to an impairment of LN LV ingrowth, resulting in a significant decrease in LN volume. These data indicate that VEGF-C/VEGFR3 signaling plays a substantial role in normal LN formation through proper migration and organization of LECs. Conclusions: Taken together, our results provide compelling evidence that the contribution of LVs through VEGF-C/VEGFR3 signaling is critical in LN development and maturation. Developmental Dynamics 245:1189-1197, 2016. (c) 2016 Wiley Periodicals, Inc.