Receptors for Short Chain Fatty Acids in the control of bacterial infection and gut immunity

The human intestine routinely contains an almost unimaginable number of bacteria. In health they perform many key roles and generate a vast number of by-products of their own metabolism that positively affect the cells and tissue of our own body. However, in certain settings invasive bacteria multiply extensively and this can result in ill-health that can range from mild symptoms of an 'upset stomach' to much more serious and life-threatening conditions such as sepsis. The gut has a surveillance system that detects the presence of such invasive bacteria, and a major element of this Gut-Associated Lymphoid Tissue is the so called 'Payer's patches'. These contain a wide range of immune cells involved in both these initial processes and then co-ordinating the response of the body to disable and destroy such bacteria. Two sets of these immune cells are called 'dendritic' cells and 'type 3 innate lymphoid (IL3)' cells. For many years we have been studying cell surface receptors that are activated by short chain fatty acids, such as acetic acid, that are generated in large amounts by the metabolic activity of many of the gut bacteria. These receptors, called Free Fatty Acid Receptor 2 and Free Fatty Acid Receptor 3 are members of a large family of transmembrane proteins called G protein-coupled receptors that are the targets of many successfully used medicines, and we have been exploring in many settings, including diabetes and other metabolic disorders, if activating or blocking one or other of these receptors for short chain fatty acids might be a useful means to treat disease. Within these studies we have developed a number of novel chemical ligands that either mimic the effects of the short chain fatty acids or block their actions. It turns out that both 'dendritic' cells and 'type 3 innate lymphoid (IL3)' cells express very high amounts of Free Fatty Acid Receptor 2 and other cells in the gut express Free Fatty Acid Receptor 3. This implies that Free Fatty Acid Receptor 2, and possibly also Free Fatty Acid Receptor 3, are likely to play important roles in the detection of invasive bacteria and promote both gut and overall whole body immune responses. A number of studies have supported this idea but other studies are contradictory.
Initial studies that are designed to understand basic biological processes and whether they can potentially be targeted to treat human disease are often performed in mice. This reflects that as mammals they share many of the same physiological functions and mechanisms of control as humans. However, there are also many differences, and in the case of Free Fatty Acid Receptor 2, potential drug-like like molecules that both we and pharmaceutical companies have developed to block this receptor are very effective at the human form of the receptor but do not work at the mouse form. As we need to fully understand the possible effects of activating or blocking this receptor in mouse before studies in human can be considered we have engineered mice so that they express the human form of the receptor in place of the mouse version. We have also made other genetic modifications to the receptor in these modified mice to allow us to 'see' clearly the cells in which the human receptor is expressed. We plan to use these mice and the variety of drug-like compounds we have developed to assess if activating the modified receptor, or indeed Free Fatty Acid Receptor 3 which is unchanged in these mice, is effective in combatting the ability of invasive bacteria including Salmonella to colonise the gut and potential cause disease. We will also use other genetic strategies to assess if activating Free Fatty Acid Receptor 2 only in dendritic' cells or 'type 3 innate lymphoid (IL3)' cells is able and sufficient to combat infection. These experiments will generate clear understanding if activating free fatty acid receptors can suggest a therapeutic strategy that can be taken forward to human studies.

G protein-coupled receptors for the short chain fatty acids (SCFAs) that are produced in copious amounts by the gut microbiota have attracted significant interest in areas at the interface of immunity and metabolism. Despite this, a major limitation in unambiguous definition of the underpinning biology and therapeutic potential of these receptors has been the limited pharmacological tool-box available with which to interrogate these receptors in rodent models of disease. In recent times we have generated a variety of such tool molecules. A further issue is that for Free Fatty Acid Receptor 2 (FFAR2), a potential target for the treatment of sepsis, available antagonists have good affinity at the human receptor but not at either the rat or mouse orthologues. To overcome this and provide suitable mouse models for study we have generated knock-in transgenic mouse lines that replace mouse FFAR2 with either the human orthologue or a DREADD-variant of FFAR2 that is not activated by SCFAs but can be activated on demand by synthetic ligands that only activate the DREADD but not wild type FFAR2 or FFAR3. Addition of an epitope tag to the introduced receptors, as well as the development of antisera that only recognise the receptor post-activation, provide a unique set of reagents with which to assess the roles of this receptor, and also FFAR3, in countering bacterial invasion and its sequalae using Salmonella and Citrobacter as models. Crossing of the FFAR2-DREADD expressing animals with lines that will limit expression to either dendritic or type 3 innate lymphoid cells will focus attention specifically on these immune cell subsets where, to support transcriptomic data, we have already shown high level expression of FFAR2 at protein level. Outcomes of these studies will support decisions on the translational potential of regulating FFAR2, FFAR3 or both these receptors for SCFAs in aspects of gut immunity and bacterial infections.