Qualitative analysis of human monocyte and macrophage proteomes after stimulation with Acanthamoeba soluble products: a preliminary study.

Acanthamoeba castellanii is a free-living amoeba ubiquitous in nature, with a worldwide distribution. Despite its free-living mode of life, Acanthamoeba can act as facultative parasite causing severe chronic infectious diseases mainly involving immune-privileged sites such as the brain and the eyes. Dissecting host innate immune system and Acanthamoeba interactions might suggest interesting insights for the development of new therapeutic regimes, addressed to ameliorate the inflammation at the site of infection while eradicating the pathogen. So far, immunological and molecular techniques have been applied to understand the mechanisms at the interface between innate immune cells and Acanthamoeba, however the proteomics approach has not been performed yet. OBJECTIVES: All this considered, the aim of the study was to characterise the proteome of human primary monocyte and macrophages after challenge with Acanthamoeba in order to understand if Acanthamoeba might modulate protein expression in these innate immune cells and the pathways are mainly involved. METHODS: Towards this purpose, the total proteome of human peripheral blood derived monocytes and human monocyte-derive macrophages, stimulated with Acanthamoeba (Genotype T4) conditioned medium (aCM), was extracted and subsequently digested for shotgun gel-free proteomic analysis through LC/MS-MS techniques. RESULTS: 3669 proteins were identified in human monocyte samples, of which 75 and 83 were found majorly expressed in un-stimulated and aCM stimulated monocytes, respectively (P<0.05, -1≤ Rsc ≥1). 4884 proteins were identified in human macrophage samples, of which 92 and 57 were found majorly expressed in un-stimulated and aCM-stimulated macrophages, respectively (P<0.05, -1≤ Rsc ≥1). Pathway analysis showed that Acanthamoeba soluble products modulated the expression of proteins involved in cell adhesion, integrin signaling pathway, inflammation mediated by chemokine and cytokine signaling pathway, antigen processing and presenting and plasminogen activating cascade in monocytes. In macrophages, stimulation with aCM modulated the expression of protein involved in complement and coagulation cascade. CONCLUSION: Overall our study provide preliminary insights into the mechanisms by which Acanthamoeba may influence protein expression in monocyte and macrophages and consequently their state of activation and the outcome of infections. The use of this newest techniques in the study of host/Acanthamoeba interactions could have practical use in pharmaceutical and diagnostic fields.