Endothelial cell polarity and extracellular matrix composition require functional ATP6AP2 during developmental and pathological angiogenesis
Nehal R. Patel, Rajan K C, Avery Blanks, Yisu Li, Minolfa C. Prieto, Stryder M. Meadows
Nehal R. Patel, Rajan K C, Avery Blanks, Yisu Li, Minolfa C. Prieto, Stryder M. Meadows
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Research Article Angiogenesis Vascular biology

Endothelial cell polarity and extracellular matrix composition require functional ATP6AP2 during developmental and pathological angiogenesis

Abstract

The (Pro)renin receptor ([P]RR), also known as ATP6AP2, is a single-transmembrane protein that is implicated in a multitude of biological processes. However, the exact role of ATP6AP2 during blood vessel development remains largely undefined. Here, we use an inducible endothelial cell–specific (EC-specific) Atp6ap2-KO mouse model to investigate the role of ATP6AP2 during both physiological and pathological angiogenesis in vivo. We observed that postnatal deletion of Atp6ap2 in ECs results in cell migration defects, loss of tip cell polarity, and subsequent impairment of retinal angiogenesis. In vitro, Atp6ap2-deficient ECs similarly displayed reduced cell migration, impaired sprouting, and defective cell polarity. Transcriptional profiling of ECs isolated from Atp6ap2 mutant mice further indicated regulatory roles in angiogenesis, cell migration, and extracellular matrix composition. Mechanistically, we provided evidence that expression of various extracellular matrix components is controlled by ATP6AP2 via the ERK pathway. Furthermore, Atp6ap2-deficient retinas exhibited reduced revascularization in an oxygen-induced retinopathy model. Collectively, our results demonstrate a critical role of ATP6AP2 as a regulator of developmental and pathological angiogenesis.

Authors

Nehal R. Patel, Rajan K C, Avery Blanks, Yisu Li, Minolfa C. Prieto, Stryder M. Meadows

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Figure 1

Defective retinal angiogenesis in EC-specific Atp6ap2-KO mice.

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Defective retinal angiogenesis in EC-specific Atp6ap2-KO mice. (A) Strat...
(A) Strategy for EC-specific deletion of Atp6ap2 in postnatal mice (Atp6ap2 induced endothelial cell KO; Atp6ap2iECKO) by tamoxifen administration at P1–P3 for early induction or at P5–P7 for late induction. Murine retinas were analyzed at indicated ages. (B) qPCR analysis of isolated lung ECs (iLECs) show Atp6ap2 expression levels in control and Atp6ap2iECKO mice at P7 (n = 3, triplicates for each sample). (C) Western blot analysis of iLECs from control and Atp6ap2iECKO mice at P7. (D) Densitometric quantification of ATP6AP2 levels in C (n = 3). (E–H) Whole-mount isolectin-IB4–stained (IB4-stained) retinas (dotted circles represent outgrowth in the control retina) (E) and quantification of radial outgrowth (F), vascular density (G), and branch points in control and Atp6ap2iECKO mice at P7 (n =7) (H). Scale bars: 1,000 µm. (I–L) Whole-mount IB4-stained retinas and closeup images of IB4+ vessels in the superficial plexus and deep plexus (I) with quantification of radial outgrowth (n = 4) (J), vascular density in the superficial plexus (n = 4) (K), and vascular density in the deep plexus (n = 4) (L) in control and Atp6ap2iECKO mice at P12. Scale bars: 1,000 µm. (M) Three-dimensional reconstructed images of the IB4+ deep vascular plexus and comparisons of the length of the perpendicular growth in control and Atp6ap2iECKO mice at P12. Note very few yellow depth-coded vessels in the deep plexus of Atp6ap2 mutants. Data are shown as mean ± SD; 2-tailed unpaired t test. **P < 0.01, ***P < 0.001, ****P < 0.0001.