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Piezo1 integration of vascular architecture with physiological force.

Li J, Hou B, Tumova S, Muraki K, Bruns A, Ludlow MJ, Sedo A, Hyman AJ, McKeown L, Young RS, Yuldasheva NY, Majeed Y, Wilson LA, Rode B, Bailey MA, Kim HR, Fu Z, Carter DA, Bilton J, Imrie H, Ajuh P, Dear TN, Cubbon RM, Kearney MT, Prasad KR, Evans PC, Ainscough JF, Beech DJ - Nature (2014)

Bottom Line: Global or endothelial-specific disruption of mouse Piezo1 profoundly disturbed the developing vasculature and was embryonic lethal within days of the heart beating.Downstream of this calcium influx there was protease activation and spatial reorganization of endothelial cells to the polarity of the applied force.The data suggest that Piezo1 channels function as pivotal integrators in vascular biology.

View Article: PubMed Central - PubMed

Affiliation: 1] School of Medicine and Multidisciplinary Cardiovascular Research Centre, University of Leeds, Leeds LS2 9JT, UK [2].

ABSTRACT
The mechanisms by which physical forces regulate endothelial cells to determine the complexities of vascular structure and function are enigmatic. Studies of sensory neurons have suggested Piezo proteins as subunits of Ca(2+)-permeable non-selective cationic channels for detection of noxious mechanical impact. Here we show Piezo1 (Fam38a) channels as sensors of frictional force (shear stress) and determinants of vascular structure in both development and adult physiology. Global or endothelial-specific disruption of mouse Piezo1 profoundly disturbed the developing vasculature and was embryonic lethal within days of the heart beating. Haploinsufficiency was not lethal but endothelial abnormality was detected in mature vessels. The importance of Piezo1 channels as sensors of blood flow was shown by Piezo1 dependence of shear-stress-evoked ionic current and calcium influx in endothelial cells and the ability of exogenous Piezo1 to confer sensitivity to shear stress on otherwise resistant cells. Downstream of this calcium influx there was protease activation and spatial reorganization of endothelial cells to the polarity of the applied force. The data suggest that Piezo1 channels function as pivotal integrators in vascular biology.

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Piezo1 mRNA in aorta and endothelial cellsa, End-point PCR products obtained with Piezo1 primers for human (h.) liver and mouse (m.) lung endothelial cell (EC) and freshly-dissected mouse aorta mRNA after reverse transcriptase reaction (RT) to generate cDNA. b, As for (a) but for: human late outgrowth endothelial progenitor cells (LEPC) and 7 types of human endothelial cell (art., arterial; micr., microvascular; pul., pulmonary; umb., umbilical; car., cardiac; bla., bladder; der., dermal; col., colonic). Results are shown with (+ RT) and without (-RT) reverse transcription. c, Quantitative real-time PCR data for experiments of the type shown in (a) (n=2 each in duplicate). d, Quantitative real-time PCR data for experiments of the type shown in (b) (n=1 each in duplicate).
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Figure 5: Piezo1 mRNA in aorta and endothelial cellsa, End-point PCR products obtained with Piezo1 primers for human (h.) liver and mouse (m.) lung endothelial cell (EC) and freshly-dissected mouse aorta mRNA after reverse transcriptase reaction (RT) to generate cDNA. b, As for (a) but for: human late outgrowth endothelial progenitor cells (LEPC) and 7 types of human endothelial cell (art., arterial; micr., microvascular; pul., pulmonary; umb., umbilical; car., cardiac; bla., bladder; der., dermal; col., colonic). Results are shown with (+ RT) and without (-RT) reverse transcription. c, Quantitative real-time PCR data for experiments of the type shown in (a) (n=2 each in duplicate). d, Quantitative real-time PCR data for experiments of the type shown in (b) (n=1 each in duplicate).

Mentions: Messenger RNA encoding Piezo1 was readily detected in mouse aorta and a variety of human endothelial cells (Extended Data Fig. 1). To gain insight into its significance we first investigated cultured human umbilical vein endothelial cells (HUVECs). Depletion of Piezo1 by either of two short interfering RNAs (siRNAs) strongly suppressed migration of these cells towards vascular endothelial growth factor (VEGF) (Extended Data Fig. 2a-e), a key stimulant of angiogenesis in vivo9. There was similar inhibitory effect of a spider toxin blocker of Piezo1 channels, GsMTx410, and a non-specific small-molecule blocker, ruthenium red6 (Extended Data Fig. 2f). Consistent with a relationship to endothelial cell migration, HUVEC tube formations in vitro and in vivo were suppressed by Piezo1 depletion (Extended Data Fig. 2g-j). We therefore generated mice with disrupted endogenous Piezo1 gene (Extended Data Fig. 3a, b). Piezo1+/− progeny appeared normal but Piezo1−/− was embryonic lethal (Fig 1a). Of 49 Piezo1−/− embryos the longest survival time was E16.5 and most were only E9.5-11.5, which is a critical time for vascular development9. At E10.5, growth retardation was commonly observed (Extended Data Fig. 3c). At E9.5, embryos were often normal in size but yolk sac vasculature was less prominent (Fig 1b). While endothelial cells were present in yolk sacs of Piezo1+/+ and Piezo1−/− embryos the structures were different, with greater disorganization and fewer defined large vessels in Piezo1−/− (Fig 1c). Similar observations were made at E10.5 (Fig 1d). The data suggest Piezo1 as a protein of critical importance in the control of vascular architecture and embryonic development.


Piezo1 integration of vascular architecture with physiological force.

Li J, Hou B, Tumova S, Muraki K, Bruns A, Ludlow MJ, Sedo A, Hyman AJ, McKeown L, Young RS, Yuldasheva NY, Majeed Y, Wilson LA, Rode B, Bailey MA, Kim HR, Fu Z, Carter DA, Bilton J, Imrie H, Ajuh P, Dear TN, Cubbon RM, Kearney MT, Prasad KR, Evans PC, Ainscough JF, Beech DJ - Nature (2014)

Piezo1 mRNA in aorta and endothelial cellsa, End-point PCR products obtained with Piezo1 primers for human (h.) liver and mouse (m.) lung endothelial cell (EC) and freshly-dissected mouse aorta mRNA after reverse transcriptase reaction (RT) to generate cDNA. b, As for (a) but for: human late outgrowth endothelial progenitor cells (LEPC) and 7 types of human endothelial cell (art., arterial; micr., microvascular; pul., pulmonary; umb., umbilical; car., cardiac; bla., bladder; der., dermal; col., colonic). Results are shown with (+ RT) and without (-RT) reverse transcription. c, Quantitative real-time PCR data for experiments of the type shown in (a) (n=2 each in duplicate). d, Quantitative real-time PCR data for experiments of the type shown in (b) (n=1 each in duplicate).
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Related In: Results  -  Collection

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Figure 5: Piezo1 mRNA in aorta and endothelial cellsa, End-point PCR products obtained with Piezo1 primers for human (h.) liver and mouse (m.) lung endothelial cell (EC) and freshly-dissected mouse aorta mRNA after reverse transcriptase reaction (RT) to generate cDNA. b, As for (a) but for: human late outgrowth endothelial progenitor cells (LEPC) and 7 types of human endothelial cell (art., arterial; micr., microvascular; pul., pulmonary; umb., umbilical; car., cardiac; bla., bladder; der., dermal; col., colonic). Results are shown with (+ RT) and without (-RT) reverse transcription. c, Quantitative real-time PCR data for experiments of the type shown in (a) (n=2 each in duplicate). d, Quantitative real-time PCR data for experiments of the type shown in (b) (n=1 each in duplicate).
Mentions: Messenger RNA encoding Piezo1 was readily detected in mouse aorta and a variety of human endothelial cells (Extended Data Fig. 1). To gain insight into its significance we first investigated cultured human umbilical vein endothelial cells (HUVECs). Depletion of Piezo1 by either of two short interfering RNAs (siRNAs) strongly suppressed migration of these cells towards vascular endothelial growth factor (VEGF) (Extended Data Fig. 2a-e), a key stimulant of angiogenesis in vivo9. There was similar inhibitory effect of a spider toxin blocker of Piezo1 channels, GsMTx410, and a non-specific small-molecule blocker, ruthenium red6 (Extended Data Fig. 2f). Consistent with a relationship to endothelial cell migration, HUVEC tube formations in vitro and in vivo were suppressed by Piezo1 depletion (Extended Data Fig. 2g-j). We therefore generated mice with disrupted endogenous Piezo1 gene (Extended Data Fig. 3a, b). Piezo1+/− progeny appeared normal but Piezo1−/− was embryonic lethal (Fig 1a). Of 49 Piezo1−/− embryos the longest survival time was E16.5 and most were only E9.5-11.5, which is a critical time for vascular development9. At E10.5, growth retardation was commonly observed (Extended Data Fig. 3c). At E9.5, embryos were often normal in size but yolk sac vasculature was less prominent (Fig 1b). While endothelial cells were present in yolk sacs of Piezo1+/+ and Piezo1−/− embryos the structures were different, with greater disorganization and fewer defined large vessels in Piezo1−/− (Fig 1c). Similar observations were made at E10.5 (Fig 1d). The data suggest Piezo1 as a protein of critical importance in the control of vascular architecture and embryonic development.

Bottom Line: Global or endothelial-specific disruption of mouse Piezo1 profoundly disturbed the developing vasculature and was embryonic lethal within days of the heart beating.Downstream of this calcium influx there was protease activation and spatial reorganization of endothelial cells to the polarity of the applied force.The data suggest that Piezo1 channels function as pivotal integrators in vascular biology.

View Article: PubMed Central - PubMed

Affiliation: 1] School of Medicine and Multidisciplinary Cardiovascular Research Centre, University of Leeds, Leeds LS2 9JT, UK [2].

ABSTRACT
The mechanisms by which physical forces regulate endothelial cells to determine the complexities of vascular structure and function are enigmatic. Studies of sensory neurons have suggested Piezo proteins as subunits of Ca(2+)-permeable non-selective cationic channels for detection of noxious mechanical impact. Here we show Piezo1 (Fam38a) channels as sensors of frictional force (shear stress) and determinants of vascular structure in both development and adult physiology. Global or endothelial-specific disruption of mouse Piezo1 profoundly disturbed the developing vasculature and was embryonic lethal within days of the heart beating. Haploinsufficiency was not lethal but endothelial abnormality was detected in mature vessels. The importance of Piezo1 channels as sensors of blood flow was shown by Piezo1 dependence of shear-stress-evoked ionic current and calcium influx in endothelial cells and the ability of exogenous Piezo1 to confer sensitivity to shear stress on otherwise resistant cells. Downstream of this calcium influx there was protease activation and spatial reorganization of endothelial cells to the polarity of the applied force. The data suggest that Piezo1 channels function as pivotal integrators in vascular biology.

Show MeSH
Related in: MedlinePlus