Extracellular Acidosis and Tissue Destruction in Tuberculosis

Tuberculosis (TB) is a leading cause of death worldwide and has been labelled a global health emergency by the World Health Organisation. TB is also a significant health problem in the UK with 10,000 new cases diagnosed each year. The current drug treatment for TB is prolonged, lasting a minimum of 6 months, which results in difficulties ensuring patients complete therapy. Resistance of TB to current drug therapy is also on the rise making TB increasingly difficult to treat. Consequently, new approaches to treating TB are required.

TB is a disease that most commonly affects the lungs but can cause disease in any part of the body. Whilst fighting the infection, the bodys' own immune system causes significant inflammation which leads to tissue destruction at the site of infection. This tissue destruction is responsible for the symptoms and mortality associated with TB as well as the spread of infection. Much of this tissue destruction is caused by production of a group of enzymes called matrix metalloproteinases (MMPs). In many infections, the site where the immune system is fighting the infection becomes more acidic than healthy tissue due to inflammation. Previous studies show that acidosis changes how the immune system responds to infection. Despite this no previous studies have ever looked at whether TB infection makes tissues acidic and laboratory studies investigating TB have never looked at how changes in acid levels could effect the immune response to TB.

To show that TB infection makes tissues acidic, we will carry out a clinical study measuring the acidity of samples taken from the lymph nodes of patients with TB of the lymph glands. With their informed consent and NHS Research Ethics Committee approval, patients will be recruited from Hammersmith, Charing Cross and Northwick Park Hospitals. We shall then carry out laboratory studies looking at how TB infected cells behave differently in acidic conditions and in particular how acidosis changes the production of MMPs and other key proteins which fight infection. This study will help us better understand how tissue destruction occurs in TB which will hopefully lead to new treatments.

The research will be conducted under the supervision of Professor Jon Friedland in the Department of Infectious Diseases and Immunity at Imperial College London.

This project investigates a new hypothesis which is that tissue acidosis arising during innate inflammatory responses to tuberculosis (TB) amplifies matrix metalloproteinase (MMP) gene expression and secretion, worsening tissue destruction. It builds on data obtained in my Academic Clinical Fellowship where I showed that acidosis increases MMP-1,3 & -9 secretion from primary human respiratory epithelial cells & macrophages independent of changes in hypoxia but potentially involving the signalling molecule AMPK. Preliminary data also showed that TB lymph nodes are acidotic. My specific aims are first to investigate effects of extracellular acidosis on MMP gene expression and secretion in TB-infected human monocyte-derived macrophages & respiratory epithelial cells and in a multicellular model of infection. Following kinetic and pH ranging experiments (pH7.4-6.7 based on initial data), I shall use luminex, ELISA, zymography and functional luminescent confocal assays to assess MMP secretion and activity. MMP qPCR will measure gene expression. Effect of acidosis on transcriptional regulators will be investigated using Trans-AM kits, siRNA and ChiP assays. Next, I shall dissect mechanisms by which acidosis regulates MMP gene expression in TB. First, I shall screen for kinase activation using mutilplex assays. I shall focus on the MAPK, PI3 kinase, and AMPK (currently being identified as key in the TB MMP response) paths. The role of the pH sensing receptor TDAG8 which activates Protein Kinase A, in the regulation of MMPs in TB will be investigated. Finally, acidosis and MMP secretion will be examined in patients with TB lymphadenitis measuring pH, MMPs and inhibitory TIMPs in aspirates and dissecting identified signalling and transcriptional control paths by immunohistochemistry. In summary, this project will provide a broad laboratory and clinical research training investigating the effects of acidosis on MMPs in TB, an area in which there is little current data.

Academic impacts:
Increased understanding of tuberculosis (TB) immunopathogenesis and in particular the mechanisms leading to tissue destruction should identify potential novel therapeutic targets. Limiting tissue damage and inflammation may help to reduce mortality and morbidity. Inflammatory sites in conditions as diverse as infection, cancer and autoimmunity have been demonstrated to be acidotic. Matrix metalloproteinases (MMPs) are also involved in the pathogenesis of these conditions and therefore the interaction between innate immunity, acidosis and MMP biology will be of interest not just to infection specialists but also to researchers studying a range of respiratory, rheumatological, renal and oncological conditions.

Economic and social impacts:
The identification of novel therapeutic targets for TB will attract pharmaceutical research and if successful, drug development. New therapies would hopefully be effective in drug resistant TB and will therefore save lives in those afflicted by this global problem. Enhanced therapies may reduce the current 6 months minimum treatment duration which would aid compliance and reduce patient follow up time. This consequently should reduce treatment costs and by improving quality of life in those of employable age, increase economic productivity. The potential social and economic benefits of improved TB treatment would be felt not just in the UK, where 10,000 new cases are diagnosed per annum, but worldwide where deaths approach 2 million a year. The timescale from commencement of the research to realisation of these more general benefits will be at least 10 years.

Training benefits:
I shall develop analytical, laboratory and information technology skills that will allow me to develop a career as an academic clinician and produce high quality research. In the future I will also use this knowledge and skills to train students, clinical fellows and postdoctoral students when I am running an independent research laboratory.

Public Engagement with Science benefits:
I will ensure that my research is disseminated in the public domain and is accessible to a lay audience. In this way it will be a small part of helping to improve the lay understanding of science. TB is already of interest to the public, as evidence by its appearance in the press, and therefore may be an important tool in this process.