WP 01 - Target Definition

1. Objectives

1. Identify new candidate biomarkers and targets for β-cell imaging in inflammation and regeneration
2. Identify new alternative spliced biomarkers for β-cell imaging
3. Validate the candidate biomarkers
4. Provide antigen structure for antibody production of the most promising candidate biomarkers
5. Set up in vitro and in vivo models for the validation of the specificity and selectivity of potential biomarkers

2.  Description

 

1. Identify new candidate biomarkers and targets for beta-cell imaging in inflammation and regeneration.
The receptome of beta cells will be identified on microarray datasets obtained in control and inflamed beta cells or islet cells (rat and human datasets) and in microarray datasets in regeneration models. We will select plasma receptors/transporters which are highly and/or selectively expressed in islets but have low expression in other intra-abdominal organs. We will also identify candidate genes that are differentially regulated during inflammation or regeneration.

2. Identify new alternative spliced biomarkers for beta-cell imaging.
Alternative splicing is responsible for much of the variation in the humane proteome (>50%). We will identify which splice variants of membrane bound proteins and thus potential biomarker candidates. The analysis will be performed on control versus inflamed islets of Langerhans and in models of regeneration. Beta cell specific receptors and their alternative splice variants will be identified.  

3. Validation of the candidate biomarkers.
In a first step, candidates will be validated at the protein level by comparing rat primary beta cells versus exocrine cells (AR42J, PANC1 cells) using Western Blotting and immuno cytochemistry. Next, expression will be tested in human islets and compared to other tissues such as liver, kidney, stomach, intestine and spleen. Subsequently the expression in in vivo models for T1D (NOD mice, MLDS injected mice), models for regeneration and transplantation will be validated.

4. Provide antigenic structure for antibody production of the most promising candidate biomarkers.
Based on the information from the identified beta cell specific receptors and their alternative splice variants expressed in beta cells (see above), we will select the extracellular antigen structure to be used for antibody production. Using IPA analysis, we will identify ligands and interacting proteins that could be labelled in WP2.

5. Set up in vitro and in vivo models for the validation of specificity and selectivity of potential biomarkers.
An in vitro model will be set up comparing rat primary beta cells, human islets cells and INS-1E cells versus AR42J or PANC-1 cells to calculate the specificity of the labelled biomarker for beta cells versus exocrine cells. Biodistribution of the labelled biomarker will be measured by injection in the tail and collection of all organs for the calculation of the tracer abundance in the different organs. In vivo models for T1D (NOD mice, multiple low dose streptozotocin (MLDS) injected mice), models for beta cell regeneration and transplantation models will be injected with labelled biomarker and analysed by imaging.  

 

3. WP leader

WP leader is Prof. Décio L. Eizirik - Université Libre de Bruxelles, Belgium

4. WP partners

Université de Genève, Switzerland
Vrije Universiteit Brussels, Belgium
University Hospital Freiburg, Germany