Canonical WNT signaling components in vascular development and barrier formation
Yulian Zhou, … , Makoto M. Taketo, Jeremy Nathans
Yulian Zhou, … , Makoto M. Taketo, Jeremy Nathans
Published August 1, 2014
Citation Information: J Clin Invest. 2014;124(9):3825-3846. https://doi.org/10.1172/JCI76431.
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Canonical WNT signaling components in vascular development and barrier formation

Abstract

Canonical WNT signaling is required for proper vascularization of the CNS during embryonic development. Here, we used mice with targeted mutations in genes encoding canonical WNT pathway members to evaluate the exact contribution of these components in CNS vascular development and in specification of the blood-brain barrier (BBB) and blood-retina barrier (BRB). We determined that vasculature in various CNS regions is differentially sensitive to perturbations in canonical WNT signaling. The closely related WNT signaling coreceptors LDL receptor–related protein 5 (LRP5) and LRP6 had redundant functions in brain vascular development and barrier maintenance; however, loss of LRP5 alone dramatically altered development of the retinal vasculature. The BBB in the cerebellum and pons/interpeduncular nuclei was highly sensitive to decrements in canonical WNT signaling, and WNT signaling was required to maintain plasticity of barrier properties in mature CNS vasculature. Brain and retinal vascular defects resulting from ablation of Norrin/Frizzled4 signaling were ameliorated by stabilizing β-catenin, while inhibition of β-catenin–dependent transcription recapitulated the vascular development and barrier defects associated with loss of receptor, coreceptor, or ligand, indicating that Norrin/Frizzled4 signaling acts predominantly through β-catenin–dependent transcriptional regulation. Together, these data strongly support a model in which identical or nearly identical canonical WNT signaling mechanisms mediate neural tube and retinal vascularization and maintain the BBB and BRB.

Authors

Yulian Zhou, Yanshu Wang, Max Tischfield, John Williams, Philip M. Smallwood, Amir Rattner, Makoto M. Taketo, Jeremy Nathans

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Figure 2

Effects of eliminating Ctnnb1 or Lrp5 and Lrp6 postnatally in ECs.

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Effects of eliminating Ctnnb1 or Lrp5 and Lrp6 postnatally in ECs. (A–O)...
(A–O) Sulfo-NHS-biotin leakage in coronal sections from WT, Lrp5–/– Lrp6CKO/CKO Pdgfb-CreER, and Ctnnb1CKO/CKO Pdgfb-CreER brains at P23–P24, following 600 μg 4HT at P15 or 200 μg 4HT at P16. Leakage is seen around the choroid plexus, meninges, and ventral hypothalamus in all 3. In the Lrp5–/– Lrp6CKO/CKO Pdgfb-CreER and Ctnnb1CKO/CKO Pdgfb-CreER brains, scattered sites of leakage are seen in ventral cortex, striatum, thalamus, paraventricular hypothalamus, pons, brainstem, and cerebellum (arrows). Scale bar: 2 mm. (P–U) Sulfo-NHS-biotin leakage in a Ctnnb1CKO/CKO Pdgfb-CreER brain at P10, following 200 μg 4HT at P7. Leakage and PLVAP and Claudin5 immunostaining are shown in paired images from cerebellum (left panels), thalamus (center panels), and cortex (right panels). Leakage was most prominent in the cerebellum, with scattered sites of leakage in cortex and anterior thalamus. The conversion of ECs from PLVAP–Claudin5+ to PLVAP+Claudin5+ or PLVAP+Claudin5– roughly correlates with sites of leakage. Arrow in U points to a vessel that expresses PLVAP. Scale bar: 200 μm. (V) Sulfo-NHS-biotin leakage in P12 (approximately) Lrp5–/– Tie2-Cre (control), Lrp5+/– Lrp6CKO/CKO Tie2-Cre, and Lrp5–/– Lrp6CKO/+ Tie2-Cre brains. There is no detectable leakage in Lrp5–/– Tie2-Cre brains (upper left) and there are rare foci of leakage (red arrows) in Lrp5+/– Lrp6CKO/CKO Tie2-Cre and Lrp5–/– Lrp6CKO/+ Tie2-Cre brains (upper panels, center and right). Lower panels show rare foci of sulfo-NHS-biotin leakage are centered on small clusters of ECs that have converted to PLVAP+Claudin5– (arrows). In the left member of each image pair, the sulfo-NHS-biotin channel has been omitted. Scale bars: 2 mm (upper panels); 200 μm (lower panels).