Neuropilin-1 deficiency in vascular smooth muscle cells is associated with hereditary hemorrhagic telangiectasia arteriovenous malformations
Sreenivasulu Kilari, Ying Wang, Avishek Singh, Rondell P. Graham, Vivek Iyer, Scott M. Thompson, Michael S. Torbenson, Debabrata Mukhopadhyay, Sanjay Misra
Sreenivasulu Kilari, Ying Wang, Avishek Singh, Rondell P. Graham, Vivek Iyer, Scott M. Thompson, Michael S. Torbenson, Debabrata Mukhopadhyay, Sanjay Misra
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Research Article Vascular biology

Neuropilin-1 deficiency in vascular smooth muscle cells is associated with hereditary hemorrhagic telangiectasia arteriovenous malformations

Abstract

Patients with hereditary hemorrhagic telangiectasia (HHT) have arteriovenous malformations (AVMs) with genetic mutations involving the activin-A receptor like type 1 (ACVRL1 or ALK1) and endoglin (ENG). Recent studies have shown that Neuropilin-1 (NRP-1) inhibits ALK1. We investigated the expression of NRP-1 in livers of patients with HHT and found that there was a significant reduction in NRP-1 in perivascular smooth muscle cells (SMCs). We used Nrp1SM22KO mice (Nrp1 was ablated in SMCs) and found hemorrhage, increased immune cell infiltration with a decrease in SMCs, and pericyte lining in lungs and liver in adult mice. Histologic examination revealed lung arteriovenous fistulas (AVFs) with enlarged liver vessels. Evaluation of the retina vessels at P5 from Nrp1SM22KO mice demonstrated dilated capillaries with a reduction of pericytes. In inflow artery of surgical AVFs from the Nrp1SM22KO versus WT mice, there was a significant decrease in Tgfb1, Eng, and Alk1 expression and phosphorylated SMAD1/5/8 (pSMAD1/5/8), with an increase in apoptosis. TGF-β1–stimulated aortic SMCs from Nrp1SM22KO versus WT mice have decreased pSMAD1/5/8 and increased apoptosis. Coimmunoprecipitation experiments revealed that NRP-1 interacts with ALK1 and ENG in SMCs. In summary, NRP-1 deletion in SMCs leads to reduced ALK1, ENG, and pSMAD1/5/8 signaling and reduced cell death associated with AVM formation.

Authors

Sreenivasulu Kilari, Ying Wang, Avishek Singh, Rondell P. Graham, Vivek Iyer, Scott M. Thompson, Michael S. Torbenson, Debabrata Mukhopadhyay, Sanjay Misra

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Figure 5

Smooth muscle cell Nrp1 deletion results in increase vascular diameter and density with a decrease in pericyte lining in the retinal vasculature.

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Smooth muscle cell Nrp1 deletion results in increase vascular diameter a...
(A) P5 retinas from Nrp1fl/fl (WT) and Nrp1fl/fl/SM22αCre+ (Nrp1SM22KO) mouse pups were stained for red isolectin-B4 (IsoB4) and NG2 for pericytes. The areas enclosed by boxes in representative images are digitally enlarged and shown as separate panels. Scale bar: 50 μm. (B) Vascular diameter was measured using Zen Pro software. Data are shown as mean diameter of artery (a), vein (v), and capillaries (cp). (C and D) Fluorescence intensity quantification of red IsoB4+ vasculature and green NG2+ (D) cells was performed using NIH ImageJ software. All images were captured at 10× objective and digitally enlarged to show as separate panels. Data are shown as mean ± SEM of n = 4–7 retinas. Nonparametric Mann-Whitney U test was performed. *P < 0.05.