Significance of Piezo1 in endothelial physiology and patho-physiology

Mechanical forces are very important for the development and maintenance of a healthy human body. This is particularly true in the cardiovascular system because of the beating of the heart and the flow of blood through blood vessels. We know, for example, that correct sensing of force by cells that line the inner wall of blood vessels (the endothelial cells) is important in protecting people against coronary artery disease, which causes heart attacks. Intriguingly, the processes by which the body senses mechanical force have remained poorly understood. Recently, however, a protein called Piezo1 was discovered. Investigating this protein may help us understand how mechanical force is sensed. This protein was discovered as a sensor of noxious mechanical hitting. That is, as a detector of mechanical insult from an external source. Our new data suggest an important and previously unrecognised perspective on Piezo1. They suggest that it is also a sensor of a more subtle physiological, internal, force call shear stress, which arises as a result of the rushing of the blood against endothelial cells. In the proposed project we will use newly-developed genetic modification of Piezo1 in mice to determine the significance of Piezo1 in vascular physiology and patho-physiology. Through this work we will provide fundamental new insight into how the cardiovascular system senses force. We think we will also provide a foundation on which new cardiovascular modulators could be developed in the future to correct or improve the way the human body senses and responds to mechanical force in disease situations. Such modulators could ultimately be useful for reducing the impact of cardiovascular disease which is the number one killer and disabler worldwide and reason for major financial burden on our societies.

The molecular mechanisms enabling endothelial cells to sense and respond to shear stress or other mechanical forces are unclear and controversial. Our unpublished/pilot data suggest that the newly-discovered Piezo1 (FAM38A) membrane protein (ion channel subunit) plays a critical and previously unrecognised role. We therefore propose to investigate the significance of Piezo1 for relevant cardiovascular parameters in vivo and to learn more about the signals upstream and downstream of Piezo1. To this end we have put in place complementary mouse genetic approaches for the in vivo manipulation of Piezo1. Using these mice we propose to apply our in vivo, ex vivo and in vitro expertise to learn about Piezo1 in the context of cardiovascular physiology and patho-physiology. Specifically we will investigate embryonic and postnatal vascular development, VEGF-induced angiogenesis in the adult mouse, and reperfusion after ischaemia. We will measure blood pressure and heart rate and characterise the in vitro contractile properties of a resistance artery. We will measure the generation of nitric oxide and reactive oxygen species because of evidence they are tightly linked to mechanical force and Piezo1. We will measure the response to mechanical injury delivered to the endothelium via a guide-wire. We will measure vascular permeability because it is linked to mechanical force and because there is evidence suggesting Piezo1 is a regulator of cell-to-cell contact. In parallel, tissues and cells from the mice will be used in ex vivo studies to deliver knowledge about downstream pathways linked to Piezo1, identified through a phosphoproteomic strategy validated in pilot studies. Factors relevant to cardiovascular physiology and disease will be tested to determine if they promote or suppress contributions of Piezo1 to mechanical sensing events in endothelial cells.

We show in this application the new information that global deletion of Piezo1 in mice is embryonic lethal. Published work has shown that point mutations in Piezo1 lead to phenotypes in humans. These observations suggest that Piezo1 is a biologically critical protein with high importance in humans and related species. We envisage therefore that the proposed Piezo1 research will provide understanding that can be used in the future to deliver new treatments or sensors to address common health problems world-wide.

According to the World Health Organisation, cardiovascular disease is the most common cause of death world-wide and a major cause of premature death and disability, leading to large financial burdens on societies, reduced life qualities, and reduced economic efficiencies. Therefore, it is important to find new and economically viable approaches to reduce the impact of cardiovascular disease. In the dominant types of cardiovascular disease, which are coronary artery disease and cerebrovascular disease, atherosclerosis is a major factor. There is consistent strong evidence in the scientific literature to suggest that endothelial cells and their responses to shear stress are critical in determining the location and severity of atherosclerotic plaques. If we can find ways to improve endothelial function and the responsiveness to mechanical strain when cardiovascular disease advances we may be able to reduce the incidence of critical cardiovascular events. Independently of atherosclerosis there may be other opportunities presented by understanding the molecular basis of mechanical sensation in the vasculature. We may, for example, be able to improve vascular responses to physical injury whether they arise through natural accident or surgical procedure. We may be able to improve perfusion of tissues because the quality of perfusion or its ability to recover from insult depends on the detection of shear stress and other mechanical forces experienced by endothelial cells through interaction with surrounding tissue. There may be therapeutic opportunities in conditions such as aneurismal disease and cancer where the mechanical force on tissues is grossly abnormal.

The proposed research is discovery research designed to reveal physiological and patho-physiological significance of a newly-discovered and intriguing protein. Understanding of this protein is expected to provide important new opportunities for delivering impact through development of devices and chemical modulators. Our team of basic scientists, cardiologists, a technologist, and a biostatistician will be well placed to provide such impact should the experiments reveal major roles of Piezo1 as we expect. We have on-going projects with medicinal chemists and chemical biologists who may be able to target extracellular domains of Piezo1 to tune mechanical sensing properties of the endothelium and improve cardiovascular outcomes.