PhD Study : The in vivo insulinotropic activity of GIP

Summary

Endogenous gut peptides, such as GLP-1 and GIP, enhance secretion of insulin induced by glucose from pancreatic β-cells¹, by elevating cytosolic levels of cAMP. The insulinotropic effect of GLP-1 agonism has been successfully confirmed both ex situ and in vivo, and stable GLP-1 mimetics have been adopted for antidiabetic clinical therapy². GIP was found less effective in diabetes patients³ and thus remains in the perspective drugs cohort, alongside with several other gut peptides, such as oxyndomodulin and xenin⁴.

The in vivo therapeutic effect of GIP could be compromised by the short lifetime of the incretin⁵ as well as the loss of the GIP-sensitivity by the target cells⁶. Whilst the former factor has been combatted successfully by using of synthetic GIP mimetics, the latter one is being researched, and so are potential additional factors modulating the GIP effect on pancreatic islet signalling. We therefore hypothesise that the lack of the in vivo effect of GIP stems from (i) the decreased responsiveness of pancreatic β-cells and (ii) the presence of additional systemic factors, limiting the impact of the long-living GIP receptor agonists.

We propose to investigate these phenomena, using the following set of experiments.

1. The acute (minutes) effect of GIP on the intracellular levels of cAMP in the islet cells in vivo. Mouse pancreatic islets will be isolated and infected with an adenovirus delivering the recombinant fluorescent cAMP sensor (‘Green Upward cADDis’, Montana Molecular, USA) in a liquid tissue culture conditions, so the sensor is expressed only in the peripheral layer of the islet cells. The islets will be then implanted into the anterior eye chamber of the recipient mouse, as outlined⁷. Upon the engraftment and vascularisation of the islets, the mouse will be immobilised and the cAMP levels inside the implanted islet cells imaged in vivo using an upright wide-field microscopic system (Olympus BX51 WI). We will specifically explore the response to a bolus injection of GIP and the novel GIP-xenin hybrid peptide ((DAla²)GIP/xenin-8-Gln)⁸, using GLP-1 and forskolin as positive controls. Adrenaline that is known to invoke differential cAMP response in pancreatic islet cells⁹, will be used as a functional marker of α-cells¹⁰.
2. We will characterise the pharmacology of the in vivo response to GIP using a battery of chemical agonists/antagonists of known signalling pathways inclusive of the inhibitors of GIP receptor that will control for the saturation of the signal by the endogenous incretin. We will furthermore explore different pre-culturing conditions for the implanted islets, such as chronic exposure to glucolipotoxic medium, depletion of β-cells with streptozotocin etc and their impact on the cAMP response to GIP.
3. We will identify genes underlying the secretory response of the islet cells to GIP in vivo, by utilising the functional labelling technology¹¹. The populations of cells displaying variations in the secretory response will be FACS-sorted for subsequent RNAseq analysis.
4. Subject to progress, we will verify our findings in the hybrid mouse-human system, by transplanting human donor islets in the mouse anterior eye chamber.

Applicants should note that Laboratory bench fees of £6,000.00 per annum are required for this self-funded PhD project.

Essential criteria

Applicants should hold, or expect to obtain, a First or Upper Second Class Honours Degree in a subject relevant to the proposed area of study.

We may also consider applications from those who hold equivalent qualifications, for example, a Lower Second Class Honours Degree plus a Master’s Degree with Distinction.

In exceptional circumstances, the University may consider a portfolio of evidence from applicants who have appropriate professional experience which is equivalent to the learning outcomes of an Honours degree in lieu of academic qualifications.

• Sound understanding of subject area as evidenced by a comprehensive research proposal
• Clearly defined research proposal detailing background, research questions, aims and methodology

Desirable Criteria

If the University receives a large number of applicants for the project, the following desirable criteria may be applied to shortlist applicants for interview.

• Completion of Masters at a level equivalent to commendation or distinction at Ulster
• Experience using research methods or other approaches relevant to the subject domain
• Sound understanding of subject area as evidenced by a comprehensive research proposal
• Work experience relevant to the proposed project
• Publications record appropriate to career stage
• Experience of presentation of research findings
• A comprehensive and articulate personal statement
• Relevant professional qualification and/or a Degree in a Health or Health related area

Funding and eligibility