Pathogenesis of Ulcerative colitis: The role of BRINP3 in colonic epithelial barrier function and mucosal inflammation

Title:
Pathogenesis of Ulcerative colitis: The role of BRINP3 in colonic epithelial barrier function and mucosal inflammation

Importance:
Ulcerative colitis (UC) is a chronic inflammatory disorder affecting 146000 people in the UK. Its treatment costs the NHS £430 millions per year to treat. It is characterised by inflammation and ulcers along the large intestine that occur intermittently causing many symptoms including diarrhoea and abdominal pain, thereby reducing the quality of life of affected individuals. It also increases risk of developing colorectal cancer by three folds and it can lead to life-threatening complications which require emergency surgery. There is no cure apart from surgical removal of the affected part of bowel and the currently available medications to reduce inflammation are associated with many serious side effects. The exact mechanism of the disease is still unknown. Thus further research to improve our understanding of the disease mechanisms is much needed so that more effective and well tolerated medical treatments can be developed and offered to patients in the future.

Our laboratory has recently discovered that an expression of a gene called BRINP3 is much lower in the bowel lining of individuals with UC compared to healthy individuals and patients with Crohn's disease indicating that reduced BRINP3 expression may be unique to UC and play an important role in disease development. BRINP3 is expressed predominately in the cells that line the colon and protect the host from the content of the bowel. These cells are also the site of disease activity and defects in their ability to maintain an effective barrier have been previously associated with UC. Currently, what function BRINP3 performs within the bowel and if reduced expression has any consequences is unknown.

To test the function of BRINP3 we have created mice that lack BRINP3 by genetic engineering. Preliminary data demonstrates that mice lacking BRINP3 have a much leakier bowel lining compared to normal mice. A leaky bowel lining has been suggested as one of the potential mechanisms responsible for the development of UC by many previous studies. Also when the BRINP3-lacking mice were treated with a chemical that induces bowel inflammation they lost much more weight and showed other markers of increased disease severity, such as diarrhoea, raised inflammatory markers (similar to what is seen in severe UC), than normal mice. These results indicate BRINP3 is playing a key role in the function of bowel and lack of it may be associated with the development of inflammation.

Through my project I will identify the role of BRINP3 at the molecular level and how lack of it changes the behavior of the cells and overall function of the bowel. I will focus my study on the permeability and inflammatory processes that seem to differ in the absence of BRINP3 by using mice that do not express BRINP3. I will also study the regulation of BRINP3 in cells and identify signalling pathways and external factors which affect gene expression. Finally, I will investigate the cellular role of BRINP3 by identifying binding partners and protein complexes that are formed within colonic cells. If I could demonstrate that BRINP3 plays a central role in maintaining mucosal integrity and a lack of it intensifies mucosal inflammation and furthermore identify a factor that increases the level of BRINP3 this can potentially be developed into a novel treatment and/or prevention of UC in future. Finally, if the lack of BRINP3 is found to be associated with increased turnover of cells this might give more insight to why patients with UC have increased risk of developing colorectal cancer.

Background:
Ulcerative colitis (UC) is a chronic inflammatory bowel disease (IBD) affecting 146000 individuals in the UK. It reduces quality of life, increases morbidities and risk of developing colorectal cancer placing significant burden on healthcare. The exact mechanism of this disease is still unknown. Complex interaction between genetic predisposition, environment factors and intestinal microbiota are believed to play role in causing immune dysregulation within the mucosa. Recent work in our laboratory has shown that there is a down-regulation of BRINP3 in colonic mucosal of UC patients compared to healthy controls. This finding was irrespective of the presence or degree of mucosal inflammation indicating that low expression of BRINP3 is unique to UC and may be playing an important role in pathogenesis. We have also shown that the low expression of BRINP3 in the mucosa is caused by increased methylation. The role of BRINP3 in humans is currently unknown.

Aims:
1. Determine the molecular and physiological role of BRINP3 in bowel mucosa and how it affects the function of the bowel mucosa and immune response in mice
2. Characterise the gene regulation and role of BRINP3 in colonic epithelial cells

Methodology:
The changes in cell proliferation, permeability and inflammation of the colonic mucosa will be examined using BRINP3 knockout mice and well characterised models of experimental colitis. The regulation of BRINP3 gene expression will be studied at the mRNA and epigenetic level. I will investigate the protein complexes and regulatory processes involving BRINP3 in colonic epithelial cells through the generation of reporter constructs, biochemistry and confocal microscopy.

Scientific and medical opportunity:
If low BRINP3 expression is shown to be playing a role in UC, an agent that increase its expression could potentially be developed as a novel treatment.

Ulcerative colitis (UC) affects more than 146000 people in the UK. Its treatment costs the NHS £430 millions per year and its incidence is increasing worldwide. It is characterised by superficial inflammation ulceration along the large intestine causing many symptoms including diarrhea, abdominal pain, and reduced quality of life in affected individuals. My research will explore the role of BRINP3 in the colonic mucosa and how it affects mucosal barrier function and regulation of inflammation. If I can show that BRINP3 is required in mucosal barrier function and reduced expression of it causes or exaggerates inflammation in UC, UC could potentially be treated or even prevented by a gene therapy that increases BRINP3 expression. This will radically alter the medical treatment of UC as currently available treatments are largely immunosuppressive therapies with many serious side effects. Thus my research could ultimately benefit patient suffering from UC, clinicians and scientists in this field, as well as researcher in fields of genetics, immunology and dysbiosis.

Also the concept of an increased mucosal permeability allowing influx of luminal antigens including bacteria to trigger immune response is believed to play a role in pathogenesis of many other autoimmune disorders such as type I diabetes and rheumatoid arthritis, another common bowel condition called irritable bowel syndrome, as well as complications of cirrhosis. Also malnutrition, which contributes to 45% of deaths amongst children under the age of 5 in developing countries, is believed to be driven by increased mucosal permeability. Therefore, investigating how the mucosal permeability is regulated by BRINP3 could contribute to a better understanding of mechanisms of these diseases leading to treatment which could potentially have an impact on millions of people globally. My findings have the potential to contribute to better quality of life, reduce mortality, reduce sick leave and increase productivity of the global society.

In the literature, significantly high or low expression of BRINP3 have been reported in several conditions such as gastric cancer, pituitary tumour and oral peri-implantitis. However, very little is known about the actual molecular and functional role of BRINP3 in humans. Thus the findings from my research will contribute to better understanding of BRINP3 in human pathophysiology in general and in these conditions.

If I could show that changing the expression of BRINP3 affects gut mucosal permeability, further research could look for a substance that regulates BRINP3 expression. Such a substance could be coupled with available medications to achieve desirable drug penetration and delivery through gut mucosa via oral or rectal administration routes thus contributing to the field of drug delivery research and pharmaceutical industry.

I will also examine how BRINP3 affects cell turnover and its potential role in the development of cancer as a defective mucosal barrier may be important in allowing luminal carcinogens to enter the tissue. Any positive findings in this regard would have an impact on the research in field of oncology.

Our preliminary data suggests BRINP3 knockout (KO) mice (that we have created) have increased mucosal permeability and have an exaggerated inflammatory response when colitis was induced chemically. Thus, researchers investigating various different diseases focusing on defective mucosal barrier function in disease pathogenesis might wish to use BRINP3 KO mice in future to conduct their experiments. Also BRINP3 mice could be established as a new mouse colitis model for future colitis research.