Modification of monocyte compartments following exposure to bacterial lysates

ABSTRACT Polyvalent mechanical bacterial lysate (PMBL) has been reported to effectively prevent and treat recurrent respiratory infections, including those of viral aetiology. It confers protection and improves immune response by modulating cellular and humoral responses. Although its immunogenic capabilities have been studied in different cell types, its fine molecular mechanisms of action remain poorly elucidated. In this study, we investigated the potential immunomodulatory effects of bacterial lysates in regulating the immune responses of human monocytes (Mo). In a well-established in vitro model of human monocytes, we found that PMBL treatment skews the conventional Mo subsets distribution, inducing a switch to a Non-classical (NC) population. We observed a sizable fraction of the Mo pool expressing high levels of surface marker CD16 with a lower level of CD14; remarkably, this subset also significantly expanded in peripheral blood after sublingual administration in individuals undergoing PMBL treatment. Expression of SLAN and CX3CR1 in the absence of CCR2, high level of HLA-DR, and preponderant expression of FcγIIIR, was the dominant pattern among the increased CD16++ cells confirming the NC-like features of this subpopulation. The transition from classical to NC subsets was observed also by culturing highly purified classical Mo with lysate, suggesting a direct involvement of PMBL in driving the conversion between the two monocyte subsets, rather than a decrease/increase of one of the two subsets PMBL could also modulate Mo immune functions leading to increased cytokine production when subsequently challenged with microbial or viral stimuli, indicating a trained immunity rewiring. Analysing their functional traits, we observed that PMBL-treated Mo displayed a boost in TNF-α, IL-6, and IL-1β response upon restimulation with LPS or R848, highlighting the role of PMBL in modulating genes of the pro-inflammatory signalling pathway, one of the main hallmarks described for the phenomenon of trained immunity. PMBL also interferes with the phosphorylation of p38-MAPK and ERK 1/2, critical intracellular signal transduction pathways involved in the inflammatory response. Additionally, PMBL training in combination with CD16 triggering resulted in a synergic effect for their effector functions. On the other hand, PMBL-treated Mo secreted a large amount of IL-10, capable of calming down and limiting hyper-responsiveness during their activation via CD16. Collectively, our results demonstrated that PMBL modulates Mo immune responses by prompting a phenotypic and functional switch toward an NC-Mo subset interfering with the intracellular signalling cascade that regulates monocyte-mediated inflammation, acquiring antibody-dependent cellular cytotoxicity (ADCC) via CD16, and endowed with increased response to the second challenge inducing trained immunity. These results indicate that PMBL could exert their protective role also through this mechanism of action. Expression of CD16 and induction of a trained immunity in Mo should enhance both innate and antigen-specific adaptive immune responses, also conferring a broader protection against different pathogens and counteracting a much wider array of diseases.