Complement activation and effector pathways in ischaemia reperfusion injury and at the interface with adaptive immunity
Ischaemia reperfusion injury (IRI) is an important source of morbidity and mortality and contributes to renal transplant failure. Renal IRI is known to be mediated by complement, which causes direct injury to the transplant and stimulates the adaptive immune response against transplant antigens. However uncertainty exists as to the exact trigger mechanisms of complement activation in this situation, and to the relative importance of certain effector molecules generated by complement activation. Based on the results of previous studies, I hypothesized that there would be greater involvement of the lectin pathway than previously recognized, as a trigger mechanism that could both initiate complement activation and lead to amplification via the alternative pathway in renal IRI. I also predicted that, whereas C5a has been shown to play a part in the genesis of IRI, C3a could be shown to have an important effect on innate cells that form a bridge with adaptive immunity. The aim of the work described in this thesis was therefore to investigate: (a) the role of the lectin pathway in IRI through the enzyme MASP-2; and (b) the role of C3a linking up the innate and adaptive immune responses following transplantation. I first investigated renal IRI in mice with deficiencies of the classical (C4), alternative (FB) and lectin (MASP-2) pathways of complement activation in order to explore the notion that the injury was mediated by the lectin pathway. Absence of C4 and FB did not protect the mice in a transplant model of IRI; but absence of MASP-2 was markedly protective suggesting that the lectin pathway - or rather a variant of it that did not involve C4 - has a significant role in the mouse kidney transplant IRI. As a result, I also investigated the therapeutic effect of anti-MASP-2 antibody in the mouse kidney isograft model. Absence of C3a-receptor in a native kidney model of IRI did not convey a substantial protective effect, suggesting no major effect of C3a in this injury. However, exploring a possible effect of C3a on the adaptive immune response, I found that stimulation of monocytes with C3a in conjunction with LPS resulted in significant IL-1β production which in turn induced Th-17 cells. I identified that the increase in monocyte IL-1β production is ATP and caspase-1 dependent and that the change in ATP is modulated by Pannexin-1 channel. My findings identify MASP-2 as an early step in the pathogenesis of complement-mediated mouse kidney IRI and support its identity as a therapeutic target. Furthermore, as both monocytes and Th-17 cells have been previously identified as major contributors to acute kidney rejection after transplantation, my results could explain, at least in part, how complement activation induced by tissue stress during the transplant procedure has an impact on the alloimmune response.
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