Chronic inflammatory diseases (CIDs) are a common group of highly disabling immune conditions leading to persistent inflammation and tissue-specific manifestations, such as ulcerative colitis and arthritis. They affect approximately 10% of the population with this figure rising as our population ages. Unfortunately treatment is expensive, rarely curative and often associated with disabling side-effects, such as a Cushingoid phenotype caused by synthetic glucocorticoid (GC) treatment. By building on excellence in inflammation and endocrinology research at the University of Birmingham, this proposal will investigate a previously ignored pathway in GC metabolism that could significantly alter our understanding of glucocorticoid metabolism in inflammatory disease.
In humans, peripheral GC metabolism, particularly through the cortisone-cortisol ‘shuttle’ via 11b-hydroxysteroid dehydrogenases, impacts inflammatory outcomes. However, both cortisone and cortisol are withdrawn from the active pool by sulfation and re-activated by desulfation through local sulfation pathways expressed in almost all tissue. However nothing is known whether sulfation pathways influence GC metabolism and action. Indeed, physiologically relevant concentrations (760-1,220 ng/100mL) of sulfated GCs are found in human plasma and most likely act as a circulating reservoir ready for desulfation by tissue.
Thus, the sulfation/desulfation activity in inflamed tissue will directly impact local bioavailability of GCs and represents an important regulator of the severity, progression, and eventual outcomes of CIDs.
Thus, we hypothesize here that:
1) GC desulfation/sulfation is a rate-limiting step to inflammatory immune responses.
2) desulfation/sulfation is dysregulated in CID with pathophysiological consequences.
Aims and Objectives
This proposal will provide fundamental new understanding on how inflammation is modified by GCs, potentially allowing for new pharmacological intervention to benefit patients with CIDs.
We will achieve this through two over-lapping work packages (WP) where we will utilise in vitro cell models, primary human inflammation models, and TNFa-Tg mice coupled with LC-MS/MS analysis to:-
1. Create enabling LC/MSMS methodologies to accurately measure sulfated GCs (WP1)
2. Provide proof-of-principle that sulfated GCs decrease inflammatory pathways (WP2).
3. Determine whether inhibition of desulfation of sulfated GCs exacerbates inflammatory outcomes in vitro (WP1+WP2)
WP1: Firstly, and with the Steroid Metabolome Analytic Core (SMAC), create new enabling methodologies for the measurement of sulphated GCs in cell media and plasma. Currently SMAC have extremely sensitive methods to measure many steroid metabolites, including cortisone and cortisol. We would aim to add to these methods the measurement of sulphated cortisone and sulphated cortisol.
WP2: Next, will be to provide proof-of-principle that sulfated GCs (cortisol, cortisone) decrease inflammation through STS activity. This will expand on our preliminary findings to underpin future in vivo work investigating the same inflammatory pathways. We will use primary human fibroblasts taken from RA patients and cell models (HCT116 cells and HepG2 cells – both significantly increase STS activity in response to TNFa (unpublished data)). We will determine whether sulfated GCs inhibit TNFa-mediated inflammation by measuring down-stream mediators (IL-1b, IL-1ra, IL-4, IL-8, IL-10, IL-17A, IFN-g, MCP-1 (MCAF) using multiplex platforms and gene array). Anti-inflammatory effects from GC conjugates will be inhibited with STX64, an irreversible specific STS inhibitor. Through designing custom-made microfluidic cards (Taqman), TNFa effects on other GC metabolism pathways (STS, SULTs, 11b-HSDs) involved in GC bioavailability will also be examined.
Significance of Research: This research will demonstrate the Proof-of-principle concept that sulfated GCs dampen-down TNFa-induced inflammation in the absence of STS inhibition. This will represent a new pathway through which GCs induce their anti-inflammatory effects and identifies sulfation pathways as potential targets to regulate anti-inflammatory effects of GCs.
Applicants should have a strong background in molecular biology, with an interest in endocrinology. They should have a commitment to anti-inflammatory research and hold or realistically expect to obtain at least an Upper Second Class Honours Degree in biology or related field.
To be considered for this studentship, please send the following documents to Dr Paul Foster [Email Address Removed]:
• A detailed CV, including your nationality and country of birth;
• Names and addresses of two referees;
• A covering letter highlighting your research experience/capabilities;
• Copies of your degree certificates with transcripts;
• Evidence of your proficiency in the English language, if applicable.