Role of the Lymphotoxin signaling axis in the regulation of thymic microenvironments: Implications for age-associated thymic atrophy.

Our susceptibility to infection gradually increases with age, directly contributing towards a negative impact upon health and wellbeing in later life. Current predictions estimate that by 2033 the proportion of individuals over the age of 65 will encompass 25% of the UK population (source: Medical Research Council, UK). It is therefore clear that tackling the problem posed by age-associated infection risk presents a major socio-economic and health challenge.

The ability of the immune system to protect against bacterial and viral infection is provided by a wide range of immune cell types acting in concert. One critical immune cell type is the T-cell, responsible for directly killing virally infected cells and supporting our ability to mount protective antibody responses. T-cells develop within a highly specialized organ, termed the thymus that lies within the upper chest cavity. Each T-cell formed within the thymus recognizes a limited range of molecules associated with specific bacteria or viruses via a cell surface receptor termed the T-cell receptor (TCR). Importantly, each T-cell produced within the body bears a single TCR specificity, thereby meaning each T-cell is capable of recognizing and attacking a limited range of infectious pathogens. In order to ensure efficient immune protection against the huge range of bacteria and viruses that an individual is likely to encounter throughout their lifetime, a diverse range of different T-cells are required to meet this diverse, infectious challenge. The central role of the thymus and T-cells in providing immune protection is acutely highlighted by the extreme risk of infection and high mortality rates in patients displaying inherited defects in thymus and T-cell development.

Throughout life, T-cells within the immune system are continuously lost through attrition. In order to ensure maintenance of T-cell diversity and efficient immune protection, continuous development and output of new T-cells from the thymus is required to counter-balance this ongoing loss of peripheral T-cells. However, as we age, the thymus undergoes a gradual program of atrophy. Thymus atrophy being characterized by: a loss of functional thymus tissue, an increase in thymus fat deposits and a corresponding restricted capacity to support new T-cell development. As a consequence, elderly individuals exhibit restricted T-cell diversity manifesting in an increased susceptibility to infection and a reduced capacity to respond to vaccination based interventions. In addition, the loss of T-cell development capacity with increasing age negatively impacts upon the ability of patients undergoing chemo- and irradiation based therapy to efficiently restore T-cell immunity post-treatment. This directly leads to a window of high susceptibility to infection following cancer-related treatments and bone-marrow transplantation.

The specialized capacity of the thymus to support T-cell development is provided by functionally unique thymic stromal cells, and it is these cells that are lost during age-associated thymus atrophy. However, it is unclear which signals and molecules are involved in either maintenance or loss of thymic stromal cells and their replacement by fibrotic tissue and fat cells in the aged thymus, cell types linked to inflammatory processes. This project will investigate the role of the Lymphotoxin-beta receptor (LTbR), a signaling molecule linked to both normal T-cell development and stromal cell function, in driving thymus atrophy. This project will also investigate the consequences of manipulating this pathway upon T-cell development and immune function. Importantly, the identification of novel molecules and their role in regulating thymus atrophy has important implications for the ultimate long-term goal of developing medical therapeutic interventions that will combat thymus atrophy and improve T-cell development and immunity in old age.

The thymus is a unique site for T-cell development, maintaining a diverse, self-tolerant T-cell pool. The ability of the thymus to support T-cell development is underpinned by specialized stromal cells. Notably, the thymus undergoes a gradual program of atrophy with increasing age. Thymic atrophy is characterized by loss of functional thymic stroma, an increase in adipocytes and fibroblasts, and a restricted capacity to support T-cell development. The functional consequence of thymus atrophy is reduced T-cell output, leading to restricted diversity within the aged peripheral T-cell pool as T-cell production rates fail to balance peripheral T-cell loss. Despite the importance of thymus atrophy to age-associated immunosenescence, the cellular and molecular pathways regulating this process are unclear. This study aims to investigate the specific role of the Lymphotoxin (LT) signalling axis during thymus atrophy. Whilst LT has been reported to play a role in organization of thymus microenvironments and thereby regulating T-cell tolerance, this mechanism remains poorly defined. Moreover, the contribution of LT signaling to perturbation in thymic stroma during thymic atrophy is currently undefined. This project will examine the role of the LT axis in the regulation of heterogeneous thymic stromal populations impacting upon thymic atrophy. Importantly, LT is implicated in the regulation of inflammatory microenvironments within the periphery, and moreover has been associated with regulating development of peripheral fat and lymphoid stroma. This project aims to define the impact of LT signalling upon: i) thymic stromal cell function and maintenance during ageing, ii) thymic fat accumulation, iii) T-cell development and tolerance using both in vitro and in vivo approaches to manipulate LT signalling. This project will provide novel data on mechanisms regulating thymic stroma and may inform future approaches to improve thymic function following age-associated thymus atrophy.

Despite the critical impact of thymus atrophy upon the increased susceptibility of aged individuals to opportunistic infection in later life, our knowledge of the cellular and molecular mechanisms that drive thymus atrophy are currently limited. The primary goal of this project is to define the role of the Lymphotoxin pathway upon age-associated alterations in thymic stromal cells that ultimately drive thymus atrophy. The results generated by this study will therefore be of interest to a wide range of end-point users interested in both understanding thymus atrophy, and also the potential for manipulation of thymus microenvironments restore T-cell function in old age. It is anticipated that the immediate beneficiaries of this research project will include basic research scientists within the field of biomedical research interested in T-cell biology and the effects of ageing on immune system function. To ensure dissemination of research findings, research data will be communicated via both Open-Access publication and presentation at research meetings. Increased impact will be ensured via sharing of methodologies through the usual scientific publication routes, and also via the use of social media. Dr Jenkinson has previously published online video protocols through video-sharing sites, including YouTube, which have provided an effective communication route for sharing novel research techniques with the wider research community. In addition to knowledge sharing, materials and data generated from aged-mouse models will potentially be shared through the Shared Ageing Research Models (ShARM) biorepository and online collaborative networking environment MiCEPACE, facilitating national and international sharing of resources in ageing research. The investigation of molecular pathways driving thymus atrophy, will potentially inform future translational approaches to boost T-cell mediated immunity in ageing populations. The study is therefore expected to be of interest to the clinical community. The impact upon human patients is predicted to be achieved via longer-term interest from pharmaceutical and drug-development industry. In addition to individual beneficiaries, the ultimate potential to translate research findings into interventional therapies may ultimately have a significant socio-economic impact due to the health and wellbeing implications of poor immune function in an increasingly aged UK population. The proposed project therefore directly addresses the BBSRC strategic priority of "Ageing Research: Lifelong Health and Wellbeing". This project will also impact in the short-term upon local scientific infrastructure and training via multiple routes. i) Regular interactions with undergraduate and postgraduate in both clinical and non-clinical education streams at the University of Birmingham provides a key opportunity to engage students at multiple career stages. It is anticipated that such engagement will expose and enthuse students to immunology and ageing research theory and methodologies. Such interactions have recently led to the recruitment and retention of high-calibre students within local PhD training schemes. ii) Professional training of the Postdoctoral Research Assistant employed for this project, and PhD students within the lab. The postdoc and students will have the opportunity to develop a broad spectrum of generic and project-specific research skills, including in vivo and ageing research techniques that align with the BBSRC Skills Statement for strategic priorities for research training. iii) Local research collaborations involving data, materials and methodology sharing, thereby contributing to the development of immunology and ageing research at Birmingham University. The project will also ensure public engagement, a major BBSRC priority, to promote dialogue with the general public, thereby communicating research outcomes and their relevance to society.