Inflammation is associated with development of atherosclerosis, and cholesterol crystals (CC) have long been recognized as a hallmark of atherosclerotic lesions. CC appear early in the atheroma ...development and trigger inflammation by NLRP3 inflammasome activation. In this study we hypothesized whether CC employ the complement system to activate inflammasome/caspase-1, leading to release of mature IL-1β, and whether complement activation regulates CC-induced cytokine production. In this study we describe that CC activated both the classical and alternative complement pathways, and C1q was found to be crucial for the activation. CC employed C5a in the release of a number of cytokines in whole blood, including IL-1β and TNF. CC induced minimal amounts of cytokines in C5-deficient whole blood, until reconstituted with C5. Furthermore, C5a and TNF in combination acted as a potent primer for CC-induced IL-1β release by increasing IL-1β transcripts. CC-induced complement activation resulted in upregulation of complement receptor 3 (CD11b/CD18), leading to phagocytosis of CC. Also, CC mounted a complement-dependent production of reactive oxygen species and active caspase-1. We conclude that CC employ the complement system to induce cytokines and activate the inflammasome/caspase-1 by regulating several cellular responses in human monocytes. In light of this, complement inhibition might be an interesting therapeutic approach for treatment of atherosclerosis.
The induction of human CD4
Th1 cells requires autocrine stimulation of the complement receptor CD46 in direct crosstalk with a CD4
T cell-intrinsic NLRP3 inflammasome. However, it is unclear whether ...human cytotoxic CD8
T cell (CTL) responses also rely on an intrinsic complement-inflammasome axis. Here we show, using CTLs from patients with CD46 deficiency or with constitutively-active NLRP3, that CD46 delivers co-stimulatory signals for optimal CTL activity by augmenting nutrient-influx and fatty acid synthesis. Surprisingly, although CTLs express NLRP3, a canonical NLRP3 inflammasome is not required for normal human CTL activity, as CTLs from patients with hyperactive NLRP3 activity function normally. These findings establish autocrine complement and CD46 activity as integral components of normal human CTL biology, and, since CD46 is only present in humans, emphasize the divergent roles of innate immune sensors between mice and men.
Cholesterol crystals (CC) are abundant in atherosclerotic plaques and promote inflammatory responses via the complement system and inflammasome activation. Cyclic oligosaccharide ...2-hydroxypropyl-β-cyclodextrin (BCD) is a compound that solubilizes lipophilic substances. Recently we have shown that BCD has an anti-inflammatory effect on CC via suppression of the inflammasome and liver X receptor activation. The putative effects of BCD on CC-induced complement activation remain unknown. In this study, we found that BCD bound to CC and reduced deposition of Igs, pattern recognition molecules, and complement factors on CC in human plasma. Furthermore, BCD decreased complement activation as measured by terminal complement complex and lowered the expression of complement receptors on monocytes in whole blood in response to CC exposure. In line with this, BCD also reduced reactive oxygen species formation caused by CC in whole blood. Furthermore, BCD attenuated the CC-induced proinflammatory cytokine responses (e.g., IL-1α, MIP-1α, TNF, IL-6, and IL-8) as well as regulated a range of CC-induced genes in human PBMC. BCD also regulated complement-related genes in human carotid plaques treated ex vivo. Formation of terminal complement complex on other complement-activating structures such as monosodium urate crystals and zymosan was not affected by BCD. These data demonstrate that BCD inhibits CC-induced inflammatory responses, which may be explained by BCD-mediated attenuation of complement activation. Thus, these findings support the potential for using BCD in treatment of atherosclerosis.
Cholesterol crystals (CC) play an essential role in the formation of atherosclerotic plaques. CC activate the classical and the alternative complement pathways, but the role of the lectin pathway is ...unknown. We hypothesized that the pattern recognition molecules (PRMs) from the lectin pathway bind CC and function as an upstream innate inflammatory signal in the pathophysiology of atherosclerosis. We investigated the binding of the PRMs mannose-binding lectin (MBL), ficolin-1, ficolin-2, and ficolin-3, the associated serine proteases, and complement activation products to CC in vitro using recombinant proteins, specific inhibitors, as well as deficient and normal sera. Additionally, we examined the deposition of ficolin-2 and MBL in human carotid plaques by immunohistochemistry and fluorescence microscopy. The results showed that the lectin pathway was activated on CC by binding of ficolin-2 and MBL in vitro, resulting in activation and deposition of complement activation products. MBL bound to CC in a calcium-dependent manner whereas ficolin-2 binding was calcium-independent. No binding was observed for ficolin-1 or ficolin-3. MBL and ficolin-2 were present in human carotid plaques, and binding of MBL to CC was confirmed in vivo by immunohistochemistry, showing localization of MBL around CC clefts. Moreover, we demonstrated that IgM, but not IgG, bound to CC in vitro and that C1q binding was facilitated by IgM. In conclusion, our study demonstrates that PRMs from the lectin pathway recognize CC and provides evidence for an important role for this pathway in the inflammatory response induced by CC in the pathophysiology of atherosclerosis.
During atherogenesis, cholesterol precipitates into cholesterol crystals (CC) in the vessel wall, which trigger plaque inflammation by activating the NACHT, LRR and PYD domains-containing protein 3 ...(NLRP3) inflammasome. We investigated the relationship between CC, complement and NLRP3 in patients with cardiovascular disease.
We analysed plasma, peripheral blood mononuclear cells (PBMC) and carotid plaques from patients with advanced atherosclerosis applying ELISAs, multiplex cytokine assay, qPCR, immunohistochemistry, and gene profiling.
Transcripts of interleukin (IL)-1beta(β) and NLRP3 were increased and correlated in PBMC from patients with acute coronary syndrome (ACS). Priming of these cells with complement factor 5a (C5a) and tumour necrosis factor (TNF) before incubation with CC resulted in increased IL-1β protein when compared to healthy controls. As opposed to healthy controls, systemic complement was significantly increased in patients with stable angina pectoris or ACS. In carotid plaques, complement C1q and C5b-9 complex accumulated around CC-clefts, and complement receptors C5aR1, C5aR2 and C3aR1 were higher in carotid plaques compared to control arteries. Priming human carotid plaques with C5a followed by CC incubation resulted in pronounced release of IL-1β, IL-18 and IL-1α. Additionally, mRNA profiling demonstrated that C5a and TNF priming followed by CC incubation upregulated plaque expression of NLRP3 inflammasome components.
We demonstrate that CC are important local- and systemic complement activators, and we reveal that the interaction between CC and complement could exert its effect by activating the NLRP3 inflammasome, thus promoting the progression of atherosclerosis.
Chronic inflammation of the arterial wall is a key element in the development of atherosclerosis, and cholesterol crystals (CC) that accumulate in plaques are associated with initiation and ...progression of the disease. We recently revealed a link between the complement system and CC-induced inflammasome caspase-1 activation, showing that the complement system is a key trigger in CC-induced inflammation. HDL exhibits cardioprotective and anti-inflammatory properties thought to explain its inverse correlation to cardiovascular risk. In this study, we sought to determine the effect of reconstituted HDL (rHDL) on CC-induced inflammation in a human whole blood model. rHDL bound to CC and inhibited the CC-induced complement activation as measured by soluble terminal C5b-9 formation and C3c deposition on the CC surface. rHDL attenuated the amount of CC-induced complement receptor 3 (CD11b/CD18) expression on monocytes and granulocytes, as well as reactive oxygen species generation. Moreover, addition of CC to whole blood resulted in release of proinflammatory cytokines that were inhibited by rHDL. Our results support and extend the notion that CC are potent triggers of inflammation, and that rHDL may have a beneficial role in controlling the CC-induced inflammatory responses by inhibiting complement deposition on the crystals.
Complement's favourite organelle—Mitochondria? Rahman, Jubayer; Singh, Parul; Merle, Nicolas S. ...
British journal of pharmacology,
July 2021, Letnik:
178, Številka:
14
Journal Article
Recenzirano
Odprti dostop
The complement system, well known for its central role in innate immunity, is currently emerging as an unexpected, cell‐autonomous, orchestrator of normal cell physiology. Specifically, an ...intracellularly active complement system—the complosome—controls key pathways of normal cell metabolism during immune cell homeostasis and effector function. So far, we know little about the exact structure and localization of intracellular complement components within and among cells. A common scheme, however, is that they operate in crosstalk with other intracellular immune sensors, such as inflammasomes, and that they impact on the activity of key subcellular compartments. Among cell compartments, mitochondria appear to have built a particularly early and strong relationship with the complosome and extracellularly active complement—not surprising in view of the strong impact of the complosome on metabolism. In this review, we will hence summarize the current knowledge about the close complosome–mitochondria relationship and also discuss key questions surrounding this novel research area.
Linked Articles
This article is part of a themed issue on Canonical and non‐canonical functions of the complement system in health and disease. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v178.14/issuetoc
•Altered-self particles such as cholesterol crystals (CC) activate complement.•CC-induced complement regulates priming, phagocytosis and NLRP3 inflammasome activation.•Complement colocalize with CC ...clefts in human carotid plaques.•The inflammatory responses to CC can be controlled by targeting complement.
In the host a diverse collection of endogenous danger signals is constantly generated consisting of waste material as protein aggregates or crystalline materials that are recognized and handled by soluble pattern recognition receptors and phagocytic cells of the innate immune system. These signals may under certain circumstances drive processes leading to adverse inflammation. One example is cholesterol crystals (CC) that accumulate in the vessel wall during early phases of atherogenesis and represent an important endogenous danger signal promoting inflammation. CC is recognized by the lectin- and classical pathways of the complement system resulting in activation of C3 and C5 with release of inflammatory mediators like the potent C5a fragment. Complement activation by CC leads to crosstalk with the NLRP3 inflammasome-caspase-1 pathway and production of IL-1β. Neutralization of IL-1β may have beneficial effects on atherosclerosis and a large clinical trial with an IL-1β inhibitor is currently in progress (the CANTOS study). However, upstream inhibition of CC-induced inflammation by using a complement inhibitor may be more efficient in treating atherosclerosis since this will block initiation of inflammation processes before downstream release of cytokines including IL-1β. Another therapeutic candidate can be broad-acting 2-hydroxypropyl-β-cyclodextrin, a compound that targets several mechanisms such as cholesterol efflux, complement gene expression, and the NLRP3 pathway. In summary, emerging evidence show that complement is a key upstream player in the pathophysiology of atherosclerosis and that therapy aiming at inhibiting complement could be effective in controlling atherosclerosis.
Abstract
Intracellular/autocrine complement proteins have emerged as critical regulators of human Th1 induction and contraction. T cells contain both intracellular C3 and C5 activation systems, with ...intracellular C3aR1 and C5aR1 stimulation driving T cell homeostatic survival and normal Th1 IFNg production, respectively. Here we demonstrate how the intracellular/autocrine C5 system is regulated by using T cells from the first described patient with C5aR2 deficiency. This patient suffers from an autoinflammatory syndrome, with enhanced inflammatory Th responses and a profound loss of naïve CD4 T cells in the blood (95% have a memory phenotype). Thus, in contrast to intracellular C5aR1 stimulation, cell surface expressed C5aR2 is an important negative regulator of inflammatory Th induction. While both C5aR1 and C5aR2 can bind C5a, we found that the carboxypeptidase-processed form of C5a, C5a-desArg, was twice as potent as C5a in reducing Th1 induction. In addition, carboxypeptidase M (CPM) expression was highly induced upon T cell activation indicating that CPM may be mediating T cell-derived C5a-desArg generation and C5aR2 stimulation. In this vein, activation of CRISPR/Cas9 generated CPMKO T cells, or of T cells in the presence of a CPM inhibitor, induced Th1 hyper-induction, which was rescued by addition of a C5aR2 agonist and reduced by adding C5a-desArg, but not C5a. The in vivo importance of T cell-expressed CPM and autocrine C5aR2 activation is demonstrated by the enhanced inflammatory Th responses in Cpm −/−mice and increased pathology caused by Cpm −/−T cells in a T cell transfer colitis model. These data highlight the importance of intracellular/autocrine C5 system regulation by CPM through C5aR2 signaling in T cell responses.
Abstract
During atherosclerosis, cholesterol precipitates into cholesterol crystals (CC) in the vessel wall which trigger plaque inflammation. In the present, study we examined the relation between ...complement, NLRP3 signalling pathways and CC in patients with coronary artery disease and carotid atherosclerosis. We analysed plasma, peripheral blood mononuclear cells (PBMC) and carotid plaques from patients with different degrees of atherosclerosis applying ELISAs, multiplex, qPCR, immunohistochemistry, gene profiling and bioinformatics. Transcripts of IL-1β and NLRP3 were increased and correlated in PBMC from patients with acute coronary syndrome. Immunohistochemistry of carotid plaques showed pronounced accumulation of C1q and terminal C5b-9 complement complex (TCC) around CC-clefts. Transcripts of C5aR1, C5aR2 and C3aR1 were increased in carotid plaques compared to control arteries. Priming of carotid plaques ex vivo with C5a and TNF followed by CC incubation resulted in pronounced upregulation of NLRP3 inflammasome components. Importantly, we found that intracellular “C5 system” is a major contributor of CC induced IL-1β in human macrophages through the action of C5aR1 present on mitochondria. Mechanisms underlaying the role of intracellular C5 in this process will be described. The data suggest that CC formation is an important extracellular and intracellular complement activator which in turn results in IL-1β expression and activation of NLRP3 in atherosclerotic plaques.
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