Complement C5a Induces Pro-inflammatory Microvesicle Shedding in Severely Injured Patients

Abstract

Initially underestimated as platelet dust, extracellular vesicles are continuously gaining interest in the field of inflammation. Various studies addressing inflammatory diseases have shown that microvesicles (MVs) originating from different cell types are systemic transport vehicles carrying distinct cargoes to modulate immune responses. In this study, we focused on the clinical setting of multiple trauma, which is characterized by activation and dysfunction of both, the fluid-phase and the cellular component of innate immunity. Given the sensitivity of neutrophils for the complement anaphylatoxin C5a, we hypothesized that increased C5a production induces alterations in MV shedding of neutrophils resulting in neutrophil dysfunction that fuels posttraumatic inflammation. In a mono-centered prospective clinical study with polytraumatized patients, we found significantly increased granulocyte-derived MVs containing the C5a receptor (C5aR1, CD88) on their surface. This finding was accompanied by a concomitant loss of C5aR1 on granulocytes indicative of an impaired cellular chemotactic and pro-inflammatory neutrophil functions. Furthermore, in vitro exposure of human neutrophils (from healthy volunteers) to C5a significantly increased MV shedding and C5aR1 loss on neutrophils, which could be blocked using the C5aR1 antagonist PMX53. Mechanistic analyses revealed that the interaction between C5aR1 signaling and the small GTPase Arf6 acts as a molecular switch for MV shedding. When neutrophil derived, C5a-induced MV were exposed to a complex ex vivo whole blood model significant pro-inflammatory properties (NADPH activity, ROS and MPO generation) of the MVs became evident. C5a-induced MVs activated resting neutrophils and significantly induced IL-6 secretion. These data suggest a novel role of the C5a-C5aR1 axis: C5a-induced MV shedding from neutrophils results in decreased C5aR1 surface expression on the one hand, on the other hand it leads to profound inflammatory signals which likely are both key drivers of the neutrophil dysfunction which is regularly observed in patients suffering from multiple traumatic injuries.

Innate Immunity Communicates Using the Language of Extracellular Microvesicles

“…This increase occurs in parallel with activation of innate immunity responses, leading to activation of the ComC and the coagulation cascade (CoaC). These observations and correlations between levels of circulating ExMVs and activation of the ComC and CoaC support an important modulatory role for ExMVs in orchestrating immune responses [3][4][5][6][7][8][9][10][11][12][13][14][15][16].…”

“…As shown in Fig. 2, innate immunity cells (i) communicate with each other by exchanging ExMVs, (ii) respond to ExMVs secreted from somatic cells (e.g., mesenchymal stromal cells or cancer cells), and (iii) send ExMV-based messages to these cells in return [3][4][5][6][7][8][9][10][11][12][13][14][15][16]. This communication is intense whenever there is activation of the immune system, as seen in pathogen infection, sterile inflammation, organ or tissue injuries, and tumor growth.…”

“…The cellular arm consists of several cell types, including neutrophils, monocytes, basophils, and eosinophils as well as mast, natural killer, dendritic, and gamma/delta T cells [1,2]. These cells respond to changes in the microenvironment caused by infectious pathogens, circulating soluble mediators, physical and metabolic stimuli, and, what is the topic of this review, the release of extracellular microvesicles (ExMVs), which are small, bilayer membrane-covered blebs circulating in biological fluids [3][4][5][6][7][8][9][10][11][12][13][14][15][16]. The innate immunity cells of the cellular arm secrete ExMVs in response to external stimuli and, along with soluble mediators, are major components of the secretome [17].…”

“…It is well established that ExMVs secreted by innate immunity cells are involved in cell-cell communication during coordinated immune responses. ExMVs also directly or indirectly modulate activities of components of the soluble arm of innate immunity [3][4][5][6][7][8][9][10][11][12][13][14][15][16]. Furthermore, they may also affect responses of the acquired adaptive immune system represented by B and T lymphocytes, contributing to regulatory crosstalk between inborn and acquired immunity [3][4][5][6][7][8][9][10][11][12][13][14][15][16][17].…”

The Functional Heterogeneity of Neutrophil-Derived Extracellular Vesicles Reflects the Status of the Parent Cell

“…PMN-EVs from cells activated by either PAF [ 97 ] or IL-8 [ 96 ] demonstrated anti-inflammatory potential by decreasing PMN recruitment [ 97 ], as well as NK cell [ 96 ] activation. C5a-induced EVs were rated as anti-inflammatory vesicles as they inhibited macrophage maturation [ 94 ]; however, a recently published work showed an opposite effect where C5a-triggered EVs activated resting neutrophils to produce ROS and induced IL-6 secretion [ 121 ].…”