Can differences in Th1 IFNg-to-IL-10 cytokine switching mediated by CD46 predict functional outcome after renal transplantation?

Kidney transplantation is the preferred treatment for kidney failure, offering a better quality of life & economic benefits compared to dialysis. However, most kidney transplants do not survive as long as recipients (approximately 10-15 years) so renal graft failure accounts for 30% of those returning to dialysis and increases the demand for organs that are already in short supply.

The commonest cause of graft failure is an immune response known as chronic rejection (CR). Surprisingly little is known about the mechanisms that cause CR. Patients with detectable antibodies released by B cells are known to be at higher risk of CR but this alone has been an insufficient explanation. Research has shown that there are a number of important immunologic factors involved including T cells, B cells, cell-signalling proteins known as cytokines and the complement system (part of the innate and adaptive immune system). There is growing evidence that it is the delicate interactions between these components that underpin CR.

T cells respond to specific pathogens within cells or cells recognised to be 'non self'. Different T cells have different functions but work together and with B cells. T helper 1 (Th1) cells are a subset of CD4+ T cells and play a major role in organ rejection or alloimmunity. Th1 cells release an inflammatory cytokine known as Interferon gamma (IFNg). However, its release is controlled by another cytokine IL-10, otherwise uncontrolled IFNg production results in severe tissue damage & death. Recent research (by co-supervisor CK) shows that the complement system also plays a critical role in the release and regulation of IFNg. This is known as the concept of 'IFNg to IL-10 switching '. Abnormalities of this switching process (either by failure to stop IFNg release or failure to produce IL-10) leads to hyperactive Th1 responses, seen in autoimmune diseases such as Rheumatoid arthritis & multiple sclerosis. There are obvious attractions to the development of therapeutic control of Th1 pathway i.e. locking cells into a desired functional state.

The primary supervisor (AD) has conducted research into the role of the B cells in CR. B cells present 'foreign' proteins known as antigens from the donated kidney to CD4+ T cells. AD has shown that the rate of functional decline is significantly faster in patients where the B cells present antigen. He has developed an assay to detect CD4+T cells responses and release of IFNg. Antigen presentation by B cells has been shown to stimulate and to suppress IFNg in vitro & immune-mediated injury to the graft. In patients where B cells activity suppresses IFNg release there is also evidence of IL-10 production. Mechanisms that regulate IFNg therefore appear to be critical determinants of transplant success.

The aim of the proposed research is to investigate 'IFNg to IL-10 switching' in alloimmunity & test the hypothesis that in patients with CR, failure of the 'IL-10 switch' is associated with functional decline, graft loss & return to dialysis. Detection, management & prevention of CR presents major challenges to transplant medicine & the understanding how the cells interact is crucial for novel strategies to promote long-term graft survival.

Chronic rejection (CR) is the biggest cause of renal allograft failure. Surprisingly little is known about the cell-mediated interactions that underpin CR.
Differences in "Interferon gamma (IFNg)-to-IL-10 switching" and contraction of Th1 pathway have been demonstrated in autoimmune disease; I will test the hypothesis that patients with CR have functional differences in antigen-specific Th1 "IFNg-to-IL-10 switching" and that these differences associate with graft function.
In HLA-typed healthy 'cones' we will demonstrate that CMV gpB antigen-specific indirect ELISPOT reactivity is associated with differences in Th1 IFNg-to-IL-10 switching using flow cytometric cytokine assay. I will attempt to identify antigen-specificity in these assays.
The microenvironment matters: autocrine secretion of IL-1B mediated by NLRP3, via C5a with C5aR1 engagement is known to suppress IL-10 production; whereas engagement with C5aR2 favours IL-10 switching. We will investigate whether signalling through C5aR1/C5aR2 correlates with IFNg or IL-10 production in our viral antigen model system.
The relevance of these findings will be assessed, by analysing patient samples from the 'OuTSMART' clinical trial to correlate Th1 switching variation with graft outcome.
The involvement of CTSL, C3, NLRP3, Il-1B and signalling through C5aRs will be confirmed in HLA-stimulated CD4+ T cells, addressing whether persistent IFNg production can be modified by manipulation of these molecular pathways.
Finally, if sequencing of C5aRs demonstrates that polymorphisms are associated with IFNg-to-IL-10 switching in CMV antigen stimulated cones; we aim to sequence C5aRs in transplant samples and look for an association between (donor antigen-specific) C5aR-polymorphism and graft outcome.
The project is expected to shed new light on the molecular basis of anti-donor T cell activity and suggest potential avenues for future therapeutic manipulation to preserve organ allograft function.

Chronic rejection, the single biggest cause of renal transplant failure is a significant clinical burden, leading to thousands of patients returning to dialysis each year. The aim of this research maximize long term graft survival, by shedding new light on the molecular mechanisms causing CR and suggesting new avenues for future therapeutic manipulation to preserve allograft function.

Renal transplantation is the treatment of choice in end-stage renal failure; transplantation improves quality of life, increases life expectancy and allows patients to return to work. It is well known that donor organs are in high demand and short supply. Thousands of people die waiting for a transplant.
Increasing graft survival by addressing CR would significantly reduce the need for need for further transplantation and reduce the numbers returning to dialysis.

Clearly further research in CR is warranted and the proposed project will contribute to the advancement of scientific knowledge, to improve health & wellbeing and ultimately the economic competitiveness of the UK. There are in fact implied benefits to all societies/countries that offer transplantation.

Eradication of CR and the need for immunosuppressive therapy is the ultimate goal. In the field of transplantation, achievement of tolerance (stable graft function in the absence of immunosuppression) is the primary ambition. The attraction is obvious; the multiple burdens of expensive immunosuppressive therapy namely life-threatening infection, diabetes, malignancy, economic costs to the NHS to name a few. There are far-reaching implications for potential beneficiaries nationally and worldwide.

There will be academic beneficiaries from those in the field transplantation both in the UK and abroad. By demonstrating differences in the regulation of T cell reactivity and associating this to meaningful trends in functional outcome will provide advances in what we understand about the mechanisms at play in CR and help to suggest areas for further research strategies for early detection, management & prevention of CR. The research into mechanisms that control Th1 cell reactivity (IL-10 switching and C5aR2 & IL-1B secretion) are in relative infancy particularly in alloimmunity (although demonstrated in autoimmunity) It is hoped that this research will significantly contribute to the current knowledge of molecular regulation of Th1 pathway in alloimmunity with repercussions across other disciplines in T cell biology, the complement system, autoimmunity, infection, malignancy and possibly beyond the field of Transplantation.