The role of sterol synthesis in promoting immune tolerance in monocytes

Abstract

Monocytes are one of the key innate immune cells that initiate an immune response in the body. However, an exaggerated immune response is linked with a condition known as sepsis, which is characterised by inflammation and other mechanisms which in severe cases can lead to organ failure. In addition to sustained, excessive inflammation, a proportion of sepsis patients are characterised by a state of immune suppression. Such a state can be described as maladapted tolerance. The mechanisms underlying immune tolerance have not been fully understood yet. However, it is understood that epigenetics and immuno-metabolic pathways play crucial roles in establishing immune tolerance. Recently, the cholesterol biosynthesis pathway has been associated with the induction of tolerance in monocytes purified from community-acquired pneumonia patients on hospitalisation. This was concluded after studying the epigenetics and gene expression networks of tolerant monocytes, which indicated the distal part of the cholesterol biosynthesis pathway, specifically sterol synthesis. Whether sterol synthesis is directly involved in establishing immune tolerance in monocytes remains unknown. To understand the importance of sterol synthesis and its relationship with immune tolerance, zaragozic acid was used as an inhibitor of sterol synthesis to test whether the monocytes would respond better to a pathogenic attack. In this study, monocytes were cultivated in an in vitro model in culture plates and differentiated into macrophages. The cells were then stimulated using LPS where PBMCs underwent two LPS stimulations and zaragozic acid was added to the cells as a treatment for each hit. The cytokine TNF-alpha levels were measured using an ELISA assay to study the macrophages' response. Moreover, Flow Cytometry was performed to examine and confirm the presence of PBMC in our samples, and to characterize the monocytes against CD14 and HLA-DR surface protein expression. After analysing the ELISA results, it was concluded that Zaragozic Acid may not influence the establishment of the tolerance phenotype.