Gender, age and oestrogen metabolism

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Priority area: Basic Bioscience Underpinning HeathKeywords: LC-MS, pulmonary circulation, oestrogen metabolism Abstract:Gender exerts profound influences on vascular health and 'healthy ageing'. Women are more at risk of developing cardio-pulmonary dysfunction and this may increase post-menopause. Few studies have, however, directly examined the possibility that gender and age may induce changes on the normal pulmonary vascular function and oestrogen metabolism that might pre-dispose women to vascular risk factors. Here we will determine gender differences in the normal function of the pulmonary vasculature, in particular the role & influence of oestrogen, oestrogen metabolism & oestrogen metabolites.Whilst there are several papers and reviews concerning the influence of oestrogen on the vasculature, the influence of oestrogen metabolites on the normal ageing vasculature is very under-researched. Likewise, the influence of gender on normal proliferative signalling pathways is largely under-investigated. Our preliminary data on human pulmonary artery smooth muscle cells (hPASMCs) suggests there are gender differences in signalling pathways & that oestrogen may be the reason for the gender differences. We have recently demonstrated that oestrogen itself can decrease signalling in hPASMCs through the BMPR2 pathway increase MAPK signalling; hence proliferation of female hPASMCs is greater than in male cells. We have shown that microRNA expression in hPASMCs can be influenced by gender and oestrogens. For example, microRNA96 is decreased in hPASMCs from female lung & this causes an increase in serotonin-induced proliferation via the 5-HT1B receptor. It is emerging that oestrogen metabolites may play a more influential role on normal vasculature function than oestrogen itself. One limitation to these investigations is our ability to actually measure oestrogen metabolism and metabolites in vascular tissue. Over the last two year we have developed a novel HPLC/LC-MS 'steroidomic' method for assessing oestrogen metabolism in hPASMCs. We can now apply this technology to understand the role of oestrogen & oestrogen metabolism in the normal function and ageing of pulmonary arteries. Year 1-2. The student would assist the development of LC-MS techniques to analyse oestrogen metabolites in hPASMCs & plasma We have already identified some metabolites that accumulate in PASMCs & that have either pro- or anti-proliferative effects & at first we will examine these (e.g. 16-OHE1/2, 4-OHE1/2, 2-OHE1/2, 2 and 4-MeOHE1/2). Following this, measurements will be made in plasma at days 7, 14, 21 and 28 of the menstrual cycle from normal healthy volunteers. Similar analysis will be made in samples from post-menopausal women and age-matched men. The student will also examine the expression of key microRNAs in these samples, especially those that may interact with oestrogen metabolism or action (e.g. miRNA-22, miRNA-206, miRNA-27b). Year 2-4. hPASMCs will be derived from healthy men & women & these will be grouped according to age. This will be in collaboration with Nick Morrell (Cambridge). The effects of normal ageing and gender on basal and stimulated oestrogen metabolism will be determined. The influence of oestrogen synthesis & metabolising enzymes on this will be determined by applying aromatase inhibitors such as anastrozole and CYP1B1 inhibitors such as TMS and /or by siRNA techniques to silence these enzymes. In addition, the activity and expression of key signalling pathways will be determined (BMPR2, pERK, pAkt, reactive oxygen species etc). The synthesis of oestrogen will be determined by examining aromatase expression & via aromatase activity assay. The student will also repeat key experiments on other pulmonary cell types such as fibroblasts & also vascular smooth muscle cells from human resistance arteries (from gluteal biopsy material). High fidelity training in in vivo skills is also available if the the student wishes this.