REPOSITORY > RESULTS

Abstract

Innate immune system acts as the first line of defence to protect the host against pathogens. The regulation of immune system and inflammatory response is crucial since dysregulated or sustained inflammation leads to pathological conditions, septic shock and tissue damage. Macrophages are innate immune cells that sense pathogens with pattern recognition receptors (PRR) on the cell membranes or in the cytosol. Recognition of pathogen- or danger-associated molecular patterns (PAMP or DAMP) triggers expression of inflammatory mediators and immune defence response genes to remove and destroy the pathogens. Inflammasomes are multi-molecular platforms that recruit and activate inflammatory caspases-1 and -11, leading to the maturation and secretion of pro-inflammatory cytokines interleukins (IL)-1β and IL-18 and pyroptotic cell death, thus contributing to innate inflammatory responses. Cysteine cathepsins are proteases involved in the intracellular protein degradation in lysosomes; however, they also participate in other (patho)physiological processes, such as apoptosis, cancer progression and invasion, immune and inflammatory response, and antigen presentation. Potentially harmful and uncontrolled proteolytic activity of cysteine cathepsins is regulated by endogenous protein inhibitors, such as cystatins. Besides their inhibitory role, cystatins have other physiological roles. However, the role of cystatins in the immune response is still not well understood.
Cystatin C is constitutively expressed and secreted from all cells with nucleus and is present in all body fluids. Cystatin C was shown to have a regulatory role in the expression and secretion of inflammatory mediators, such as nitric oxide, from immune cells upon stimulation with lipopolysaccharide (LPS) or interferon (IFN)-γ. Therefore, the aim of our work was to elucidate the role of cystatin C in the inflammatory response. We examined the role of cystatin C in the inflammasome activation, release of inflammatory cytokines, innate immune signalling pathways, release of nitric oxide and autophagy induction. We demonstrated that cystatin C-deficient mice are more sensitive to LPS-induced sepsis. Further, cystatin C-deficient macrophages secrete higher amounts of pro-inflammatory cytokines IL-1β and IL-18 due to the increased caspase-1 and -11 activation. The increase in the processing and secretion of IL-1β in cystatin C-deficient macrophages was independent of the broad spectrum cathepsin inhibitor, indicating that increased cathepsin activity was not responsible for excessive inflammatory response in the absence of cystatin C. Cystatin C deficiency had no effect on NF-κB and MAPK signalling pathways, lysosomal membrane integrity, reactive oxygen species production and mitochondrial damage. We have shown that cystatin C-deficient macrophages present dysfunctional autophagy with a reduced level of autophagosomes and decreased autophagy induction upon LPS stimulation. Autophagy has an important modulatory role in inflammation. Our data suggest that the decreased autophagy in the absence of cystatin C leads to excessive inflammatory response, resulting in the increased sensitivity of cystatin C-deficient mice to the LPS-induced sepsis.