IL-6 Receptor Inhibition by Tocilizumab Attenuated Expression of C5a Receptor 1 and 2 in Non-ST-Elevation Myocardial Infarction

Abstract

Background: Elevated interleukin-6 (IL-6) and complement activation are associated with detrimental effects of inflammation in coronary artery disease (CAD). The complement anaphylatoxins C5a and C3a interact with their receptors; the highly inflammatory C5aR1, and the C5aR2 and C3aR. We evaluated the effect of the IL-6 receptor (IL-6R)-antagonist tocilizumab on the expression of the anaphylatoxin receptors in whole blood from non-ST-elevation myocardial infarction (NSTEMI) patients. Separately, anaphylatoxin receptor expression in peripheral blood mononuclear cells (PBMC) from patients with different entities of CAD was investigated.Materials and Methods: NSTEMI patients were randomized to one dose of tocilizumab (n = 28) or placebo (n = 32) and observed for 6 months. Whole blood samples drawn at inclusion, at day 2, 3 and after 6 months were used for mRNA isolation. Plasma was prepared for analysis of complement activation measured as sC5b-9 by ELISA. Furthermore, patients with different CAD entities comprising stable angina pectoris (SAP, n = 22), non-ST-elevation acute coronary syndrome (NSTE-ACS, n = 21) and ST-elevation myocardial infarction (STEMI, n = 20) were included. PBMC was isolated from blood samples obtained at admission to hospital and mRNA isolated. Anaphylatoxin-receptor-expression was analyzed with qPCR using mRNA from whole blood and PBMC, respectively.Results: Our main findings were (i) Tocilizumab decreased C5aR1 and C5aR2 mRNA expression significantly (p < 0.001) and substantially (>50%) at day 2 and 3, whereas C3aR expression was unaffected. (ii) Tocilizumab did not affect complement activation. (iii) In analyzes of different CAD entities, C5aR1 expression was significantly increased in all CAD subgroups compared to controls with the highest level in the STEMI patients (p < 0.001). For C5aR2 and C3aR the expression compared to controls were more moderate with increased expression of C5aR2 in the STEMI group (p < 0.05) and C3aR in the NSTE-ACS group (p < 0.05).Conclusion: Expression of C5aR1 and C5aR2 in whole blood was significantly attenuated by IL-6R-inhibition in NSTEMI patients. These receptors were significantly upregulated in PBMC CAD patients with particularly high levels of C5aR1 in STEMI patients.

In response to complement anaphylatoxin peptides C3a and C5a, human vascular endothelial cells migrate and mediate the activation of B‐cells and polarization of T‐cells

“…Relative levels of RNA transcripts of C3aR (A) and C5aR1 38 are shown as determined from RNA extracts isolated from endothelial cells stimulated with purified C3a or C5a, in comparison to untreated controls. Values are represented as mean ± SE analyzed from three independent experiments run in duplicate test wells, and samples duplicated for the quantitative RT‐PCR analyses (N = 12) 38 . Western blot analysis of activated C3aR (c‐Jun and ETS1) and C5aR1 (NFYA1) transcription factors from HUVECs stimulated with C3a or C5a.…”

“…Western blot analysis of activated C3aR (c‐Jun and ETS1) and C5aR1 (NFYA1) transcription factors from HUVECs stimulated with C3a or C5a. Phosphorylated (activated) pETS1 and pc‐Jun are calculated and depicted in the bar graphs of 38 as phosphorylated proteins/total proteins (see Section 2 for details). Values are represented as mean ± SE analyzed from two independent experiments run in duplicate test wells of the SDS‐PAGE gels (N = 4).…”

“…Active NFYA1 is calculated and the data shown as a bar graph of the ratio of NFYA1 to α/β tubulin. Values are represented as mean ± SE analyzed from two independent experiments run in duplicate test wells of the SDS‐PAGE gels (N = 4) 38 . Representative images of three independent experiments of HUVECs grown on cover slips that were unstimulated (negative controls) or stimulated with C3a (100 nM) or C5a (100 nM) and analyzed by immunofluorescence staining of C3aR (PE labeled), C5aR1 (PE labeled), CD31 (FITC labeled), and cell nuclei (DAPI) 57 .…”

Covid‐19: Time for a paradigm change

“…More recently, studies of coronary artery disease (CAD) found that IL‐6 and complement may both contribute to the progression of cardiovascular diseases. In patients with non‐ST‐elevation myocardial infarction (NSTEMI), IL‐6 inhibition with the anti‐IL‐6R mAb (tocilizumab) reduced C5aR1 and C5aR2 in whole blood along with decrease of C‐reactive protein (CRP) and percutaneous coronary intervention (PCI)‐related troponin T (TnT) release 13 …”

Neuregulin-4, an Adipokine, as a Residual Risk Factor of Atherosclerotic Coronary Artery Disease

“…In addition, the drugs that affect the adipokines are also being examined as potential therapeutics. [2][3][4][5][6][7][8] A recently identified adipokine, neuregulin-4 (Nrg4), is one of the four neuregulin family proteins: cell-cell signaling proteins that possess an EGF-like domain and bind to the ErbB family tyrosine kinase receptors. 9) Nrg4 was first identified as a neuregulin family protein enriched in the pancreas, 10) but recently Rosell, et al reported that Nrg4 is highly expressed in brown adipose tissues (BAT) as an adipokine; its role in atherosclerosis has been gaining attention.…”