The use of MPO as a target to treat DCI after SAH would herald a paradigm shift in types of therapies for DCI. Previous treatment strategies focused on preventing vasospasm associated with DCI6,7. More than physical disability, patients with DCI have cognitive deficits3,5,22. In this study, we show that a small molecule MPO inhibitor, AZD5904 (designated MPOi), prevents the late development of memory deficits that models change in mental status that occurs when patients develop DCI. In this model, late memory deficits are long-standing, similar to findings in human DCI18. Based on this data, small molecule MPO inhibition administered outside the blood brain barrier is a viable target for clinical trials in patients with SAH.
Inflammation as a cause of DCI after SAH has become a more popular theory in the last 20 years23–25. Unfortunately, the term inflammation is extremely broad and encompasses many pathways as different as early non-specific innate immunity and adaptive immunity with long-term immunological memory. Understanding not just that inflammation is a causative agent but also, what specific pathways are involved is critical. Here, we have focused on a subset of the innate immune system, the neutrophil response. Neutrophils, traditionally thought to be early, short-lived destructive cells that enter areas of infection and inflammation, have been found to do a multitude of other ‘jobs’ in the right environment ranging from phagocytosis to antigen presentation and cytokine release (reviewed in 26). These functions also call into question the early actor designation. In SAH, neutrophils enter the CNS days after the initial hemorrhage and do not appear to act as destructive cells19,20.
This study adds to the understanding of myeloperoxidase in the development of DCI after SAH. We have previously shown that the infiltration of neutrophils into the meninges after SAH is a critical step in the development of DCI18,19. The role of MPO in normal function and disease has evolved (reviewed in 27). MPO catalyzes the reaction between superoxide and halide ions to form halogenating agents such as hypochlorite. It was originally identified for its role in early infection. Neutrophil extracellular traps (NETs), chromatin extrusions decorated with MPO and other enzymes, play a role in a diverse set of infectious and non-infectious diseases28. In addition, there has been interest in non-enzymatic functions of MPO, including autocrine and paracrine activation of neutrophils27. This study supports the activation function of MPO in SAH. Meningeal infiltration is a critical step in the development of DCI in murine SAH. Depletion of neutrophils systemically, MPO null mice, and inhibition of the nuclear element RAR-related orphan receptor 𝛾t (ROR 𝛾t) prevents infiltration of neutrophils into the meninges and downstream complications such as vasospasm and late memory deficits18–20. In this study, we show that in the same model of SAH, small molecule inhibition of MPO improves late long-term memory deficits caused by subarachnoid hemorrhage. Importantly, MPO inhibition also prevents neutrophil entry into the meninges which is consistent with the hypothesized mechanism that neutrophil entry into the meninges is MPO mediated and a critical step.
How neutrophils in the meninges lead to dysfunction of hippocampal circuits that gate late, long-term potentiation is not understood. It is also not understood whether the action of MPOi is due to the prevention of neutrophils entering the meninges. There is much left to study, but this study shows that it is feasible in this model to prevent long-term spatial memory loss by inhibiting MPO.
A strength of this work is that it more completely characterizes the behavioral response by SAH mice than previous works in this model. We were interested not only in the time it took animals to find the goal box, but also why there was a latency. Several factors were investigated: speed the animal moved around the table, distance the animal travelled to get to the goal box, and the ‘strategy’ the animal used to find the box. Only speed to the goal box was different between the four groups. More work is needed to better understand what memory functions are altered in SAH mice. There also appear to be sex differences between SAH and sham that need further investigation.
All animal model experiments have limitations. First are variables such as model effect and center effects that could make this study less generalizable29,30. Second, we used a range of doses of AZD5904 for technical reasons. Although we found signals at all doses tested, there is still no clarity about the optimal dose of the drug. Third, we have a limited understanding of how memory losses in mice recapitulate the patient experience in SAH. Cognitive studies of SAH patients suggest that memory function is impaired although true movement-modulated spatial memory is not tested in neuropsychological evaluations. Only multicenter animal trials using multiple models and human testing will answer these questions.