The Effect of Remote Ischaemic Preconditioning on the Immune Response

Remote ischaemic preconditioning (RIPC) describes the phenomenon where brief intermittent periods of limb ischaemia are used to protect the heart and other organs from subsequent prolonged ischaemic insults. RIPC has been identified as a promising intervention for use during cardiac surgery and has consistently shown a beneficial effect in animal models; however, the results of early clinical trials have not been as successful. The exact mechanisms involved in mediating RIPC have not yet been characterised and a better understanding of the pathways through which RIPC exerts its protective effects will be essential in order to progress the translation of this intervention into the clinical setting. There is increasing evidence that RIPC modifies the inflammatory response, therefore the central aim of the research presented in this thesis was to investigate how RIPC affects the human immune system.  We performed a double-blind randomised controlled trial of RIPC in 96 high-risk cardiac surgery patients and found no evidence that the intervention reduced myocardial injury or altered peri-operative expression levels of the key inflammatory cytokines, interleukin (IL)-6, IL-8, and IL-10, during simple or more complex procedures. There was a trend towards higher levels of IL-6 and IL-8 in the preconditioned patients; however, confounding variables in the trial design and the heterogeneous patient population limited our ability to interpret the results.  We next conducted a paired-analysis trial with 10 healthy male volunteers to assess the direct effect of preconditioning on the early immune response, away from any form of ischaemic injury or comorbidities. We found that RIPC directly and significantly decreased serum levels of the chemokines MIP-1α and MIP-1β, but did not increase the serum concentrations of a range of key cytokines or alter the cytokine producing potential of peripheral blood leukocytes. These findings strongly suggest that a cytokine is not likely to be the humoral mediator associated with transmitting the RIPC protective signal.  RIPC did not alter the immunophenotype or extravasation of peripheral leukocyte populations, or the proliferative and cytokine responses of peripheral blood mononuclear cells (PBMC) to pharmacological, physiological, and antigen-specific stimuli. However, preconditioning did appear to reduce the ability of monocytes and neutrophils to respond to activation signals, as indicated by lower levels of CD11b expression in stimulated cultures, and a significant increase in the basal production of IL-22 was also detected in PBMC cultured for 6 days following preconditioning. These alterations may reduce neutrophil and monocyte tissue infiltration and limit the inflammatory response during the early window of RIPC-induced protection and enhance tissue and wound repair several days later. A multivariate analysis confirmed that there was a significant difference in the response between the control and RIPC treatments and the main contributing factors were identified as changes in neutrophil and T cell activation, serum levels of MIP-1α and β, and production of IL-10 and IL-22 from PBMC cultured for 6 days.  Overall, our results suggest that RIPC has a subtle but direct effect on the systemic innate immune response during the early window of protection in healthy volunteers, whereas the effects on the adaptive immune system seem to be considerably delayed. The changes detected following RIPC are likely to contribute to protection against ischaemia-reperfusion injury but not solely account for the extent of the beneficial effects of RIPC detected in animals. Our findings reinforce the safety profile of this intervention and have defined a number of immune parameters that are altered by preconditioning for focusing future research.