Red Cell Physical Properties in Health and Disease

Lead Research Organisation: University of Exeter
Department Name: Physics

Abstract

The red blood cell membrane has remarkable physical properties, which are major determinants of blood flow, particularly in the microcirculation. For example, blood flow through small capillaries requires fine-tuning of membrane shape and elasticity. Many diseases are associated with impaired microvascular function which has been attributed to changes in the physical properties of the red cell membrane arising from the oxidative stress to which the cell is subjected. More recently, strong evidence has begun to appear that the RBC also plays an active role in controlling microcirculation through the release of chemical signalling agents. For example, release of ATP from red cells (which stimulates the dilation of blood vessels), is stimulated by mechanical deformation of the red cell membrane, and is compromised in diabetes. Such evidence makes clear that the physical properties of red, and other, cells play important roles in controlling biological function.

In this project we will develop new experimental methodologies to analyse quantitatively the physical properties of the red cell membrane on the basis of novel computational models of membrane dynamics. We will investigate, experimentally and theoretically, the role different lipid species play in setting membrane elasticity and membrane electrical properties, how different lipid species are organised in larger formations called lipid microdomains, and how these processes are affected by oxidative stress. With the help of these insights, we will develop new computational models of the cell as a whole which, in combination with novel experimental methodologies, will enable us to accurately measure the elastic constants of the red cell. We will use these novel methodologies in the the final part of the project to investigate how the red cell membrane physical properties (elasticity, electrical potentials) affect biochemical sugnalling originating from the red cell. In these studies, we shall seek to establish whether membrane deformation, mechanical properties and/or oxidative stress affect production and release of two signalling agents, ATP (a known vesodilator) and sphingosine-1-phosphate (a signalling sphingolipid and a regulator of the vascular and immune systems).

We expect that these studies will contribute to the understanding of the origin of membrane elasticity in red cells, how it is changed in disease an how it can affect biochemical signalling pathways, as well as suggest new therapeutic targets and approaches.

Planned Impact

This project is expected to lead to a number of academic and societal impacts. Although this is not a clinical study, we expect impacts to arise in the area of healthcare and future healthcare technologies. There is overwhelming experimental evidence that oxidative stress contributes critically to the development and detrimental effects of many diseases, posing an urgent and currently unfilled need for accessible methods to monitor damage caused by oxidative stress and responses to therapeutic measures. We propose that the red cell could be used as a reporter for disease and its progression if our hypothesis that RBC physical properties correlate with clinical markers for disease is confirmed. This would offer opportunities for development of novel and unconventional technologies for assessing the effect of oxidative stress and new monitoring tools for disease progression based on quick single cell measurements. Such an approach will be of relevance to the pharmaceutical industry as well, as it will provide a novel screening mechanism to establish possible antiglycation/antioxidative properties of new drugs which may lead to a wider spectrum of applications for existing formulations.

This is a highly interdisciplinary project and will contribute towards developing and strengthening the role biophysics plays in addressing fundamental disease-related problems. Biophysics is currently one of the most dynamic areas of science and has the potential to contribute not only to the understanding of normal physiology and biological function, but also significantly add to the understanding of impaired function in disease. In the process, we will also train highly skilled interdisciplinary researchers whose future work is likely to focus on the boundary between physics, biology and medicine, either in academia or outside it, and thus contribute to the wider society.

We would also like to engage in increasing public understanding of inter- and multidisciplinary science, in particular biophysics. This will give us the opportunity to raise awareness specifically of the contribution of physics to biology and medicine. So far, physics has largely been perceived as contributing to biology and healthcare through instrument development, but our aim will be to highlight the numerous new original approaches biophysics can offer in unravelling biological function in both health and disease.

Publications

10 25 50
 
Description The membrane of living cells have a complex molecular composition and structure, and contain hundreds of different types of lipid molecules. Usually, lipid molecules are capable of assembling in microscopic domains in the membranes, which are vital for the proper function of the membrane and the whole cell. The full impact of this complexity on the membrane properties and biological function are yet to be comprehensively understood. In this project, we wet out to investigate in detail the molecular organisation of the membrane, and how it is reflected in membrane physical properties and structure. The second main aim of the project was to understand how oxidative stress, ubiquitous in many different diseases, affects biological membranes. The main findings are, as follows:
(1) We found that the different molecular composition between the inner and outer leaflet of biological membranes regulate their elasticity and microdomain structure. This research was carried out at three different synchrotron facilities in the UK, France and Germany. The results were explained using computational methods, in which the whole membrane or one of its leaflet is simulated theoretically, which allows to understand the role of each type of lilid molecules in the overall membrane organisation.
(2) We have investigated in detail what effects oxidative stress has on the structure and physical properties of membranes similar to the membrane of living cells. We discovered that oxidation of cholesterol (one of the most important molecules in the membrane) changes the membrane structure which in turn may affect membrane biological function. This research was only prossible to undertake at major international synchrotron facilities, Diamond Light Source in the UK, the European Synchrotron Radiation Facility in France, and the Germal Synchrotron Facility DESY. In order to explain these important findings, we used comutational methods to simulate the same types of membranes. We also established how the physical properties of the membrane (its elasticity and fluidity) are changed as a result of oxidation of cholesterol, one of the most important molecules in biological membranes.
(3) We also investigated the effect of DMSO on biological membranes. DMSO is an important compound used in cryopreservation of cells, but it is not entirely clear whether and how DMSO can affect biological membranes. We established that DMSO affetcs the elasticity of the cell membrane. More importantly, small amounts of DMSO cause leakage of ATP from the inside to the outside of the cell. These results are important since DMSO is used for example for preservation (banking) of umbilical cord blood rich in progenitor and stem cells. Decrease of ATP content in the preserved cells caused by DMSO may influence the cell viability and functionality upon thawing, since many cell functions dependend on energy provided by ATP. The results obtained by us may be significant in devising new cell cryopreservation strategies.
(4) We also investigated the effect of oxidative stress on the membrane of human red blood cells. We demonstrated that different oxidants make the membrane stiffer and, very importantly, impair the ability of the red cell to release ATP molecules upon deformation. Release of ATP by red cell is an important process, since ATP is involved in the regulation of the vascular tone. Our work suggests that oxidative stress, which leads to stiffening of the cell membrane, may be partially responsible for vascular complication in diseases characterised by high levels of reactive oxigen species.
Exploitation Route The results of this work will be of interest to biomedical scientists, as it may have implications about the normal and abnormal physiology of the red cell.
Sectors Healthcare

 
Description This research was set within the wider context aimed at developing fundamental biophysical approaches and methods which are capable of shedding light on impaired biological function in disease. We wanted to promote multidisciplinary approaches to address this wide and complex problem, as it clearly cannot be fully understood within the scope of a single discipline. To reach a wider cross-disciplinary impact, in April 2019 we organised an international workshop on Membranes in Health and Disease (https://nms.kcl.ac.uk/lorenz.lab/wp/?page_id=1355), attended, and contributed to, by scientists of different backgrounds (biophysicists, medical scientists, chemists, biologists, theoretical physicists and computer scientists). Due to the success of this event, two of the team members (PGP and CPW) extended the impact by acting as Guest Editors for a special issue of the journal Membranes entitled Membrane Biological Function in Health and Disease (https://www.mdpi.com/journal/membranes/special_issues/membrane_health_disease). We believe this will draw many more scientists from different backgrounds to address impaired cell membrane functions from the point of view of fundamental biophysics. Our work has also contributed to widening of the public understanding of science. Striking fluorescence microscopy images obtained by us have been disseminated through different channels, such as the Science as Art Gallery at the Exeter Science Centre (https://exetersciencecentre.org/gallery/peter_bob_uoe_lipids_both/), one of the main channels for popularising scientific research in the South West, as well as their use in promotional material to popularise research at the University of Exeter.
First Year Of Impact 2019
Sector Education,Healthcare
Impact Types Cultural,Societal

 
Description EPSRC DTP
Amount £70,810 (GBP)
Organisation Engineering and Physical Sciences Research Council (EPSRC) 
Sector Public
Country United Kingdom
Start 08/2016 
End 03/2020
 
Description Structure and mechanical properties of lipid membranes 
Organisation University of Perugia
Country Italy 
Sector Academic/University 
PI Contribution In the framework of this collaboration, we investigate the effect of dimethyl sulphoxide (DMSO) on the mechanical and structural properties of model membranes and the cell plasma membrane. The Exeter research team contributed with expertise and carried experimental work to establish quantitatively the effect of DMSO, at varying concentration, to the elastic properties of the red cell plasma membrane. We also demonstrated, for the first time, that DMSO, even at relatively low concentrations of 3%, is capable of increasing the plasma membrane permeability to ATP. These results could be of physiological importance as it is a common practice to use DMSO in relatively large concentrations (10%) as a cryoprotectant, for example for preservation (banking) of umbilical cord blood rich in progenitor and stem cells. Decrease of cell ATP levels caused by DMSO may influence the cell viability and functionality upon thawing, since a large number of cell functions and properties are dependent on ATP levels. The results obtained by us may be significant in devising new cell cryopreservation strategies. Under this collaboration, a PhD student from the University of Perugia spent several months at the University of Exeter. During this period, she undertook experiments to investigate the effect of DMSO on the membrane mechanical properties of giant lipid vesicles, as part of this project. This was made possible by the expertise of the Exeter team on membrane mechanics.
Collaborator Contribution The Perugia group have carried out extensive investigation of the effect of DMSO on the molecular structure of artificial bilayer lipid membranes using Fourier Transform Infrared spectroscopy. It was established that DMSO affects membranes differently depending on their composition and/or physical state. This is significant, because biological membranes are thought to be laterally inhomogeneous, consisting of microdomains of different composition and physical properties (lipid rafts). DMSO is threfore likely to have a differential effect on the heterogeneous membrane depending on its local composition and structure.
Impact B. Gironi, Z. Kahveci, B. McGill, B.-D. Lechner, S. Pagliara, J. Metz, A. Morresi, F. Palombo, P. Sassi* and P. G. Petrov*, Effect of DMSO on the mechanical and structural properties of model and biological membranes, Biophysical Journal, 119 (2019) 274-286.
Start Year 2018
 
Description One day interdisciplinary workshop on Membranes in Health and Disease 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact The aim of the workshop was to bring together researchers and practitioners working on physical, biological and medical aspects of cell membranes in order to initiate and sustain longer-term multidisciplinary collaborations addressing fundamental and translational questions of membranes in normal physiology and disease. The workshop created an informal atmosphere for fruitful exchanges between biophysicists, computational physicists, computer scientists and cell and molecular biologists. We are aware of further collabotarions which were seeded at the workshop.

In particular, a collaboration was initiated between our laboratory at Exeter and Dr Lesley Bruce from the Component Development Laboratory (NHS Blood and Transplant) to investigate the properties of reticulocytes from one of their cell lines. The collaboration started in 2019 and is active currently.

Workshop attendants have established connections with the Red cell Special Interest Group at the British Blood Transfusion Society (https://www.bbts.org.uk/whatwedo/sigs/redcell/) and plan to attend their annual meeting.
Year(s) Of Engagement Activity 2019