Inhibitor of apoptosis proteins (IAPs) expression in monocyte to macrophage differentiation and M1/M2 polarization

Resumen

Monocytes and macrophages constitute the first line of defense of the immune system against external pathogens and fulfill other essential roles in tissue homeostasis. The ability of monocytes to differentiate into macrophages constitutes an important asset in inflammatory situations because it allows for the quick recruitment of immune cells from the bloodstream to the site of infection. Furthermore, macrophages are highly plastic and can respond differently to changes in their environment and acquire a more cytotoxic and pro-inflammatory phenotype (M1) or a more healing and anti-inflammatory one (M2) and perform different roles during inflammation and resolution of inflammation. This polarization capacity enables the innate immune system to quickly respond to threatening situations faster than tissue specific differentiation or adaptive immune responses. Both processes, differentiation and polarization of macrophages, are important in inflammation resolution and their deregulation promotes the development of several pathologies, including cancer. On the other hand, macrophage activation syndrome (MAS), is a life-threatening complication of some rheumatic diseases, it is caused by an excessive activation and expansion of macrophages and T-lymphocytes. The inhibitor of apoptosis protein family (IAPs) has important roles in the regulation of several cellular processes and in innate and adaptive immune responses, specifically in many macrophage functions and almost every signaling pathway leading to NF-κB activation, which is implicated in pro-inflammatory cytokine production. In this thesis we wanted to characterize the expression profile of IAPs in the differentiation of monocytes to macrophages and in polarization into M1/M2 states, study the effect of IAPs pharmacological inhibition in macrophage differentiation and polarization and analyze the possibility of IAPs targeting as a therapeutic approach for macro- phage activation syndrome. We first set and optimized the in vitro macrophage differentiation and polarization models for the human monocytic cell lines THP-1 and U937 or primary monocytes from human peripheral blood mononuclear cells. We exposed them to different treatments (PMA for macrophage differentiation, LPS and IFN-γ for M1 and IL-4 for M2 macrophages) and, once we established the models, we analyzed the characteristic expression pattern of some members of the IAP family, namely, NAIP, cIAP1 and cIAP2 during monocyte to macrophage differentiation and in macrophage polarization. We have also evaluated the impact of pharmacological inhibition of IAPs in macrophage differentiation and polarization using a SMAC mimetic compound (SMC), SMC-LC161, an IAP antagonist currently used in cancer clinical trials. We also used SMC-LC161 to analyze the basal expression of cIAP1 and cIAP2 in MAS and the effect of IAP antagonism in a MAS-like mouse model. We have seen that NAIP decreases its expression in the monocyte to macrophage differentiation process but increases when macrophages are polarized to M2, which we hypothesize is related to the implication of NAIP in cell proliferation. cIAP1 and cIAP2 present an inverse pattern of expression in polarized macrophages, with especially elevated levels of cIAP2 in M1 macrophages. Interestingly, the SMAC mimetic treatment leaded to an upregulation of NAIP, principally in M2, at the same time promoted the reduction of cIAP1 in M1 and M2 and a substantial increase of cIAP2 in M1. In the MAS-like mouse model we found that the treatment with SMC-LCL161 is not a valid approach to treating the disease, as the intended therapy did not revert the characteristic MAS features. In addition, SMC treatment of monocyte derived macrophages from MAS patients did not show a significant alteration in cIAP1 or cIAP2 expression after treatment. These findings may lead to a better understanding of macrophage function and may contribute to the development of therapies in inflammatory conditions based on polarization switching.