Cyclodextrin Reduces Cholesterol Crystal-Induced Inflammation by Modulating Complement Activation.

Cholesterol crystals (CC) are abundant in atherosclerotic plaques and promote inflammatory responses via the complement system and inflammasome activation. Cyclic oligosaccharide 2-hydroxypropyl-ß-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.

TLR8 Senses Staphylococcus aureus RNA in Human Primary Monocytes and Macrophages and Induces IFN-ß Production via a TAK1-IKKß-IRF5 Signaling Pathway.

Staphylococcus aureus may cause serious infections and is one of the most lethal and common causes of sepsis. TLR2 has been described as the main pattern recognition receptor that senses S. aureus and elicits production of proinflammatory cytokines via MyD88 -: NF-κB signaling. S. aureus can also induce the production of IFN-ß, a cytokine that requires IFN regulatory factors (IRFs) for its transcription, but the signaling mechanism for IFN-ß induction by S. aureus are unclear. Surprisingly, we demonstrate that activation of TLR2 by lipoproteins does not contribute to IFN-ß production but instead can suppress the induction of IFN-ß in human primary monocytes and monocyte-derived macrophages. The production of IFN-ß was induced by TLR8-mediated sensing of S. aureus RNA, which triggered IRF5 nuclear accumulation, and this could be antagonized by concomitant TLR2 signaling. The TLR8-mediated activation of IRF5 was dependent on TAK1 and IκB kinase (IKK)ß, which thus reveals a physiological role of the recently described IRF5-activating function of IKKß. TLR8 -: IRF5 signaling was necessary for induction of IFN-ß and IL-12 by S. aureus, and it also contributed to the induction of TNF. In conclusion, our study demonstrates a physiological role of TLR8 in the sensing of entire S. aureus in human primary phagocytes, including the induction of IFN-ß and IL-12 production via a TAK1 -: IKKß -: IRF5 pathway that can be inhibited by TLR2 signaling.

Reconstituted High-Density Lipoprotein Attenuates Cholesterol Crystal-Induced Inflammatory Responses by Reducing Complement Activation.

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.

Cholesterol crystals induce complement-dependent inflammasome activation and cytokine release.

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-1ß, 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-1ß 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-1ß release by increasing IL-1ß 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.