Dataset Information

Blood Vessel Patterning on Retinal Astrocytes Requires Endothelial Flt-1 (VEGFR-1).

ABSTRACT: Feedback mechanisms are critical components of many pro-angiogenic signaling pathways that keep vessel growth within a functional range. The Vascular Endothelial Growth Factor-A (VEGF-A) pathway utilizes the decoy VEGF-A receptor Flt-1 to provide negative feedback regulation of VEGF-A signaling. In this study, we investigated how the genetic loss of flt-1 differentially affects the branching complexity of vascular networks in tissues despite similar effects on endothelial sprouting. We selectively ablated flt-1 in the post-natal retina and found that maximum induction of flt-1 loss resulted in alterations in endothelial sprouting and filopodial extension, ultimately yielding hyper-branched networks in the absence of changes in retinal astrocyte architecture. The mosaic deletion of flt-1 revealed that sprouting endothelial cells flanked by flt-1-/- regions of vasculature more extensively associated with underlying astrocytes and exhibited aberrant sprouting, independent of the tip cell genotype. Overall, our data support a model in which tissue patterning features, such as retinal astrocytes, integrate with flt-1-regulated angiogenic molecular and cellular mechanisms to yield optimal vessel patterning for a given tissue.

SUBMITTER: Chappell JC

PROVIDER: S-EPMC6787756 | biostudies-literature | 2019 Sep

REPOSITORIES: biostudies-literature

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Blood Vessel Patterning on Retinal Astrocytes Requires Endothelial Flt-1 (VEGFR-1).

Chappell John C JC Darden Jordan J Payne Laura Beth LB Fink Kathryn K Bautch Victoria L VL

Journal of developmental biology 20190907 3

Feedback mechanisms are critical components of many pro-angiogenic signaling pathways that keep vessel growth within a functional range. The Vascular Endothelial Growth Factor-A (VEGF-A) pathway utilizes the decoy VEGF-A receptor Flt-1 to provide negative feedback regulation of VEGF-A signaling. In this study, we investigated how the genetic loss of <i>flt-1</i> differentially affects the branching complexity of vascular networks in tissues despite similar effects on endothelial sprouting. We se ...[more]

PMID: 31500294

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Project description:BackgroundAngiogenesis, or the growth of new vasculature from existing blood vessels, is widely considered a primary hallmark of cancer progression. When a tumor is small, diffusion is sufficient to receive essential nutrients; however, as the tumor grows, a vascular supply is needed to deliver oxygen and nutrients into the increasing mass. Several anti-angiogenic cancer therapies target VEGF and the receptor VEGFR-2, which are major promoters of blood vessel development. Unfortunately, many of these cancer treatments fail to completely stop angiogenesis in the tumor microenvironment (TME). Since these therapies focus on the biochemical activation of VEGFR-2 via VEGF ligand binding, we propose that mechanical cues, particularly those found in the TME, may be a source of VEGFR-2 activation that promotes growth of blood vessel networks even in the presence of VEGF and VEGFR-2 inhibitors.ResultsIn this paper, we analyzed phosphorylation patterns of VEGFR-2, particularly at Y1054/Y1059 and Y1214, stimulated via either VEGF or biomechanical stimulation in the form of tensile strains. Our results show prolonged and enhanced activation at both Y1054/Y1059 and Y1214 residues when endothelial cells were stimulated with strain, VEGF, or a combination of both. We also analyzed Src expression, which is downstream of VEGFR-2 and can be activated through strain or the presence of VEGF. Finally, we used fibrin gels and microfluidic devices as 3D microtissue models to simulate the TME. We determined that regions of mechanical strain promoted increased vessel growth, even with VEGFR-2 inhibition through SU5416.ConclusionsOverall, understanding both the effects that biomechanical and biochemical stimuli have on VEGFR-2 activation and angiogenesis is an important factor in developing effective anti-angiogenic therapies. This paper shows that VEGFR-2 can be mechanically activated through strain, which likely contributes to increased angiogenesis in the TME. These proof-of-concept studies show that small molecular inhibitors of VEGFR-2 do not fully prevent angiogenesis in 3D TME models when mechanical strains are introduced.

Project description:BackgroundPericytes and vascular smooth muscle cells, collectively known as mural cells, are recruited through PDGFB (platelet-derived growth factor B)-PDGFRB (platelet-derived growth factor receptor beta) signaling. MCs are essential for vascular integrity, and their loss has been associated with numerous diseases. Most of this knowledge is based on studies in which MCs are insufficiently recruited or fully absent upon inducible ablation. In contrast, little is known about the physiological consequences that result from impairment of specific MC functions. Here, we characterize the role of the transcription factor SRF (serum response factor) in MCs and study its function in developmental and pathological contexts.MethodsWe generated a mouse model of MC-specific inducible Srf gene deletion and studied its consequences during retinal angiogenesis using RNA-sequencing, immunohistology, in vivo live imaging, and in vitro techniques.ResultsBy postnatal day 6, pericytes lacking SRF were morphologically abnormal and failed to properly comigrate with angiogenic sprouts. As a consequence, pericyte-deficient vessels at the retinal sprouting front became dilated and leaky. By postnatal day 12, also the vascular smooth muscle cells had lost SRF, which coincided with the formation of pathological arteriovenous shunts. Mechanistically, we show that PDGFB-dependent SRF activation is mediated via MRTF (myocardin-related transcription factor) cofactors. We further show that MRTF-SRF signaling promotes pathological pericyte activation during ischemic retinopathy. RNA-sequencing, immunohistology, in vivo live imaging, and in vitro experiments demonstrated that SRF regulates expression of contractile SMC proteins essential to maintain the vascular tone.ConclusionsSRF is crucial for distinct functions in pericytes and vascular smooth muscle cells. SRF directs pericyte migration downstream of PDGFRB signaling and mediates pathological pericyte activation during ischemic retinopathy. In vascular smooth muscle cells, SRF is essential for expression of the contractile machinery, and its deletion triggers formation of arteriovenous shunts. These essential roles in physiological and pathological contexts provide a rationale for novel therapeutic approaches through targeting SRF activity in MCs.

Dataset Information

Blood Vessel Patterning on Retinal Astrocytes Requires Endothelial Flt-1 (VEGFR-1).

Publications

Blood Vessel Patterning on Retinal Astrocytes Requires Endothelial Flt-1 (VEGFR-1).

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