MRC Precision Medicine DTP: Estrogenic profiling in lung in Pulmonary Arterial Hypertension by Mass Spectrometry Imaging

Background
Pulmonary Arterial Hypertension (PAH) is a progressive and deadly disease leading to right heart failure, lacking effective treatment. Remodeling and narrowing of the blood vessels in the lung is caused by accelerated proliferation of fibroblasts, smooth muscle and endothelial cells. The disease affects more women than men, suggesting that aberrant estrogen signaling may be causative. Mutations in bone morphogenetic protein receptor 2 (BMPR2) cause heritable PAH and estrogen signalling may modify this pathway in idiopathic disease, although whether this mechanism is unified throughout patients is unknown1. Through genome screening, MacLean has identified key enzymes transforming estradiol and estrone (reproductive estrogens) into proliferative metabolites (e.g. 16-hydroxylated estrogens) and recently with Andrew has demonstrated that these steroids are present in higher amounts in patients. Thus key estrogen metabolising enzymes CYP1B1 and CYP1A12 may be novel therapeutic targets for PAH and CYP1A1 antagonists are of interest.

Toxic estrogen metabolites must now be measured within larger populations and also directly within lung. Andrew and MacLean have developed mass spectrometry (MS) analysis to evaluate the estrogen profiles (a panel of 10 bioactive steroids) and gained access to valuable patient cohorts. The project will explore relationships between aberrant estrogen metabolism in different types of the disease e.g. idiopathic vs heritable and also between male and female patients, where prevailing estrogen levels will differ substantially. Furthermore, estrogen metabolism specifically in lungs must be assessed to ascertain where the global changes in estrogen metabolic profiles are reflected in the lung, the primary site of disease.

Hypothesis: Estrogen metabolism by CYP1A1 in PAH increases damaging proliferative metabolites within lung.
1) The student will examine estrogen metabolism in human primary cell models of PAH to assess if the different forms (genetic, idiopathic) of disease have the same biochemical fingerprint and also to compare sexes. Pulmonary vascular cells will be cultured and incubated with estradiol and metabolic profiles in cell medium assessed. Analysis will be performed by LC-MS/MS.
2) The estrogen profile in lung will be assessed regionally in rodent models of PAH (SUGEN treated rat). We will apply MS imaging, a technique which allows regional sampling from surfaces. The metabolites desorbed are assigned specific co-ordinates and distribution maps superimposed with histological changes. This will provide unique insight into the local environment of disease. The student will develop approaches to chemically tag estrogenic metabolites on lung surfaces, similar to approaches for androgen and glucocorticoids developed by Andrew).
3) MS imaging will also be used to gain insights into wider sterol/lipidomic changes in lungs affected by PAH. Oxidative stress in PAH promotes changes in oxidised lipids demonstrated in plasma of affected patients. Using the unique capacity of spatial lipidomics, we will evaluate lipidomic changes in lungs, again using the SUGEN rat model.

Training outcomes:
The student will train in bioanalytical MS, merging expertise in chemistry and biology. They will learn how to co-register 3D datasets with other imaging modalities e.g. histology and learn molecular biological and protein chemistry techniques as necessary to identify cell-types. They will undergo training in in vivo skills and learn how to interrogate bioinformatic lipidomics databases, involving programming skills and how to conduct multivariate statistics of omics datasets. The project will operate at the interface of preclinical and clinical medicine and the student will have the opportunity to gain from training programmes in both Edinburgh and Glasgow. They will operate in an environment in Edinburgh modelled on GLP standards, raising their awareness of industrial standards.
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This MRC programme is joint between the Universities of Edinburgh and Glasgow. You will be registered at the host institution of the primary supervisor detailed in your project selection.

All applications should be made via the University of Edinburgh, irrespective of project location:

http://www.ed.ac.uk/studying/postgraduate/degrees/index.php?r=site/view&id=919

Please note, you must apply to one of the projects and you should contact the primary supervisor prior to making your application. Additional information on the application process if available from the link above.

For more information about Precision Medicine visit:

http://www.ed.ac.uk/usher/precision-medicine