Mechanism underlying the development of Crohn's disease: the role of optineurin in macrophages
Lead Research Organisation:
University College London
Department Name: Medicine
Abstract
Crohn‘s disease is a chronic inflammatory condition of the gut that causes significant ill health and loss of earning. Earlier studies we performed showed that patients with Crohn‘s disease were less able to clear bacteria from their tissues. This is due to defective release of antimicrobial compounds by immune cells resulting from an abnormal expression of proteins that transport compounds inside the cell.
A recent screen has found abnormal production of the protein optineurin in 8 patients (13%) with Crohn‘s disease. Optineurin is a prime candidate for causing defective release of antimicrobial compounds because it plays a role in the transport of compounds for release from a cell.
The aim of this project is to determine the function of optineurin in antimicrobial defence and its role in causing Crohn‘s disease. I will be looking at the optineurin gene to identify mutations. I will also find out where optineurin is expressed in the gut and how optineurin works normally and when mutated.
I hope that by finding out the function of optineurin in antimicrobial defence and how it directs the transport of compounds inside the cell, we can determine its role in causing Crohn‘s disease so treatment targets can be developed.
A recent screen has found abnormal production of the protein optineurin in 8 patients (13%) with Crohn‘s disease. Optineurin is a prime candidate for causing defective release of antimicrobial compounds because it plays a role in the transport of compounds for release from a cell.
The aim of this project is to determine the function of optineurin in antimicrobial defence and its role in causing Crohn‘s disease. I will be looking at the optineurin gene to identify mutations. I will also find out where optineurin is expressed in the gut and how optineurin works normally and when mutated.
I hope that by finding out the function of optineurin in antimicrobial defence and how it directs the transport of compounds inside the cell, we can determine its role in causing Crohn‘s disease so treatment targets can be developed.
Technical Summary
Background
Crohn‘s disease (CD) is a chronic inflammatory condition of the gastrointestinal (GI) tract that affects approximately 85,000 individuals in the United Kingdom and causes significant morbidity and loss of earning. Genome-wide association studies have identified genes that suggest an aberrant interaction between bowel contents and the immune system resulting in CD.
Recent work in our laboratory revealed a common phenotype for CD with impaired bacterial clearance due to decreased neutrophil accumulation. The latter is likely related to abnormal macrophage function because of decreased pro-inflammatory cytokine secretion required for neutrophil chemotaxis. This was not due to reduced gene transcription but to targeted intracellular destruction of cytokine precursors in lysosomal compartments. A clue to the underlying mechanism for this was provided by the differential transcription profiles for genes encoding vesicular trafficking proteins in macrophages. These findings echo recent genome-wide association studies in CD that have identified associations with vesicular trafficking proteins.
A subsequent microarray study on CD patients analysed using outlier analysis software revealed a number of abnormally expressed molecules in macrophages. The most striking was optineurin (OPTN), which is involved in vesicular trafficking. OPTN was decreased in 8 CD patients (13%). OPTN interacts with vesicular trafficking proteins such as huntingtin, Rab8, and myosin VI, which are involved in post-Golgi trafficking to lysosomes and vesicle fusion events.
Aims
I hypothesise that OPTN plays a central role in the pathogenesis of CD via defects in vesicular trafficking and cytokine secretion in macrophages resulting in decreased neutrophil accumulation and bacterial clearance. I aim to:
1. Determine the DNA sequence, expression and distribution of OPTN in monocytes, macrophages and the GI tract of CD patients.
2. Define the mechanism by which OPTN is involved in the secretion of cytokines by macrophages when stimulated by bacteria.
Methodology
Ethical approval has been granted to study patients with CD at University College London and Royal Free Hospitals.
The OPTN promoter and exons will be resequenced. OPTN expression in the GI tract will be determined using immunohistochemistry and laser capture microdissection of GI macrophages from bowel biopsies. To define the functional effects of wild-type and mutant OPTN in macrophage cytokine secretion, immunoprecipitation, confocal microscopy and molecular cloning techniques will be utilised.
Scientific and medical opportunities
I hope to identify mutations in OPTN and define the mechanisms causing defective vesicular trafficking and cytokine secretion in macrophages. This will enable potential therapeutic targets and drugs to be developed for CD.
Crohn‘s disease (CD) is a chronic inflammatory condition of the gastrointestinal (GI) tract that affects approximately 85,000 individuals in the United Kingdom and causes significant morbidity and loss of earning. Genome-wide association studies have identified genes that suggest an aberrant interaction between bowel contents and the immune system resulting in CD.
Recent work in our laboratory revealed a common phenotype for CD with impaired bacterial clearance due to decreased neutrophil accumulation. The latter is likely related to abnormal macrophage function because of decreased pro-inflammatory cytokine secretion required for neutrophil chemotaxis. This was not due to reduced gene transcription but to targeted intracellular destruction of cytokine precursors in lysosomal compartments. A clue to the underlying mechanism for this was provided by the differential transcription profiles for genes encoding vesicular trafficking proteins in macrophages. These findings echo recent genome-wide association studies in CD that have identified associations with vesicular trafficking proteins.
A subsequent microarray study on CD patients analysed using outlier analysis software revealed a number of abnormally expressed molecules in macrophages. The most striking was optineurin (OPTN), which is involved in vesicular trafficking. OPTN was decreased in 8 CD patients (13%). OPTN interacts with vesicular trafficking proteins such as huntingtin, Rab8, and myosin VI, which are involved in post-Golgi trafficking to lysosomes and vesicle fusion events.
Aims
I hypothesise that OPTN plays a central role in the pathogenesis of CD via defects in vesicular trafficking and cytokine secretion in macrophages resulting in decreased neutrophil accumulation and bacterial clearance. I aim to:
1. Determine the DNA sequence, expression and distribution of OPTN in monocytes, macrophages and the GI tract of CD patients.
2. Define the mechanism by which OPTN is involved in the secretion of cytokines by macrophages when stimulated by bacteria.
Methodology
Ethical approval has been granted to study patients with CD at University College London and Royal Free Hospitals.
The OPTN promoter and exons will be resequenced. OPTN expression in the GI tract will be determined using immunohistochemistry and laser capture microdissection of GI macrophages from bowel biopsies. To define the functional effects of wild-type and mutant OPTN in macrophage cytokine secretion, immunoprecipitation, confocal microscopy and molecular cloning techniques will be utilised.
Scientific and medical opportunities
I hope to identify mutations in OPTN and define the mechanisms causing defective vesicular trafficking and cytokine secretion in macrophages. This will enable potential therapeutic targets and drugs to be developed for CD.
