Dissecting the development and localisation of protective IL-10-secreting T cells in a model of hepatic immunopathology

Many chronic diseases in humans, including type-1 diabetes, irritable bowel syndrome and alcohol-induced liver cirrhosis are due to the development of inappropriate and excessive inflammation in vulnerable organs of the body. Much of the pathology associated with infection is also due to over-activation of the host's own immune system. Inflammation-associated pathology is also a common cause of ill health and loss of productivity in companion animal and livestock species, with implications for animal welfare and food security. Organ and tissue damage caused by the body's own immune system is called immunopathology and is, in general, due to accumulation and activation of immune cells (leukocytes) within the inflamed organ. It is clear that before we can develop better therapies to ameliorate immunopathology, we need a much better understanding of how 'healthy' inflammatory responses are regulated and why these protective regulatory responses sometimes fail. Inflammation of the liver can lead to acute liver failure or, if chronic, to cirrhosis and eventually, liver failure. Common causes of liver failure include infection (e.g. hepatitis virus infections) and alcohol abuse, as well as automimune diseases and cancer. We aim to examine the development of protective (regulatory) immune responses in a mouse model of acute liver immunopathology. We will focus on a specific population of leukocytes (called CD4+ T cells) that produce the soluble mediator, interleukin-10 (IL-10). It is becoming clear that IL-10 producing CD4+ T cells are important for controlling inflammation in many different organs, including the liver but, at present, we do not know where these cells develop, which cells they develop from, what the signals are that cause them to differentiate or how they accumulate in inflamed organs. To answer these questions we will generate a colony of mice in which the cells that cause the inflammation (effector cells) produce a yellow fluorescent protein and the IL-10-secreting cells produce a green fluorescent protein, so that we can identify the different cell types as they differentiate and migrate to different organs during the development of liver disease and so that we can observe the changes in the cell populations when we use various experimental approaches to either induce or block their development. The results of this work will reveal how we can induce, or inhibit, the development of IL-10 producing cells and how we can enhance or reduce their accumulation in particular organs in order to prevent pathology. This should assist in the development of new drugs to ameliorate immunopathology in the liver and in other organs.

Failure of tissue homeostasis due to excessive inflammation during infection or in autoimmune conditions frequently leads to immunopathology which, if not treated, may cause organ failure and, in severe cases, death. Understanding how the immune system attempts to limit tissue-specific inflammation is essential if we are to develop effective therapies to prevent or reverse inflammation-induced organ damage. Accumulating evidence suggests that IL-10 producing CD4+ T cells (CD4+IL-10+ T cells) are crucial for the prevention of pathology in a range of distinct inflammatory conditions. Nevertheless, at present we have a very limited understanding of how these cells accumulate within non-lymphoid tissues during inflammation. We will use a model of hepatic inflammation (acute murine malaria) and a novel IFN-gamma and IL-10 dual reporter system (yellow and green fluorescence) to investigate in vivo the pathways controlling the accumulation of CD4+IL-10+ T cells in inflamed tissues. In particular we will determine (i) whether IL-10-producing cells differentiate in lymphoid tissues and then migrate to sites of inflammation or whether they differentiate in situ in the inflamed liver and (ii) whether they differentiate directly from naïve T cells or by reprogramming of effector cells. Using a variety of techniques including cell sorting, immunocytochemistry, intravital 2-photon imaging and in vitro co-culture, we will characterise effector (IFN-gamma+)and IL-10+ CD4 T cell responses in the spleen, the inflamed liver and its draining lymph nodes and determine the molecular signals required for their differentiation and migration. Our results will provide important information regarding the ontology of tissue IL-10+ CD4 T cells and the essential signals for their effective accumulation at sites of inflammation. This information should assist in the development of new drugs to ameliorate immunopathology in the liver and in other organs

This study will provide essential information that should facilitate the development of novel anti-inflammatory drugs and biologicals. Our data should provide new leads for development of novel products that may be used as treatments for acute or chronic inflammatory disorders. Perhaps more importantly, if we can identify pathways leading to accelerated IL-10 responses, or to long term retention of IL-10 secreting cells at sites of chronic inflammation, we may open up avenues for interventions that completely prevent the development of immune-mediated damage or that effectively cure inflammatory diseases rather than just managing the consequences of inflammation. Thus, a major impact of our work is likely to be on the biotechnology and pharmaceutical industry. Although we will provide specific information on liver disease, our findings are likely to be relevant for many other forms of inflammation. Our work should therefore have a direct impact on academic researchers working on many other inflammatory disorders. Our data should also provide new leads for people researching the molecular mechanisms of T cell subset differentiation, migration, tissue localisation and ontogeny. In addition, the dual reporter mouse will be of use to many of these basic researchers.