SARS-CoV-2 spike protein promotes IL-6 trans-signaling by activation of angiotensin II receptor signaling in epithelial cells

Abstract

Cytokine storm is suggested as one of the major pathological characteristics of SARS-CoV-2 infection, although the mechanism for initiation of a hyper-inflammatory response, and multi-organ damage from viral infection is poorly understood. In this virus-cell interaction study, we observed that SARS-CoV-2 infection or viral spike protein expression alone inhibited angiotensin converting enzyme-2 (ACE2) receptor protein expression. The spike protein promoted an angiotensin II type 1 receptor (AT1) mediated signaling cascade, induced the transcriptional regulatory molecules NF-κB and AP-1/c-Fos via MAPK activation, and increased IL-6 release. SARS-CoV-2 infected patient sera contained elevated levels of IL-6 and soluble IL-6R. Up-regulated AT1 receptor signaling also influenced the release of extracellular soluble IL-6R by the induction of the ADAM-17 protease. Use of the AT1 receptor antagonist, Candesartan cilexetil, resulted in down-regulation of IL-6/soluble IL-6R release in spike expressing cells. Phosphorylation of STAT3 at the Tyr705 residue plays an important role as a transcriptional inducer for SOCS3 and MCP-1 expression. Further study indicated that inhibition of STAT3 Tyr705 phosphorylation in SARS-CoV-2 infected and viral spike protein expressing epithelial cells did not induce SOCS3 and MCP-1 expression. Introduction of culture supernatant from SARS-CoV-2 spike expressing cells on a model human liver endothelial Cell line (TMNK-1), where transmembrane IL-6R is poorly expressed, resulted in the induction of STAT3 Tyr705 phosphorylation as well as MCP-1 expression. In conclusion, our results indicated that the presence of SARS-CoV-2 spike protein in epithelial cells promotes IL-6 trans-signaling by activation of the AT1 axis to initiate coordination of a hyper-inflammatory response.

Experimental data using candesartan and captopril indicate no double-edged sword effect in COVID-19

“…We analyzed TNF-α, IL-6 and CCL2. TNF-α [ 60 ] and IL-6 [ 32 , 61 ] have been identified as major inducers of the SARS-CoV-2 hyperinflammatory response. CCL2 is considered a fibrosis-associated chemokine involved in several lung inflammatory disorders including SARS, asthma and pulmonary fibrosis.…”

“…Several mechanisms may be involved in the changes in proinflammatory cytokine release observed in the present study. First, we observed a spike-induced down-regulation of the anti-inflammatory RAS axis and up-regulation of the proinflammatory AT1/NOX2 axis, which may lead to an increase in cytokine release [ 32 , 63 , 64 ] mediated by NOX2-derived ROS [ 54 , 65 ]. Second, ADAM17 is not only involved in shedding of ACE2 [ 49–51 ], but in the release several cytokines [ 66 ].…”

“…Second, ADAM17 is not only involved in shedding of ACE2 [49][50][51], but in the release several cytokines [66]. It is well-known that ADAM17 or TACE (TNF-α -converting enzyme) releases TNF-α from cells [67], and that ADAM17 promotes IL-6/soluble IL-6 receptor release [32]. In addition, ADAM17 may induce CCL2 release via an indirect mechanism involving acidic mammalian chitinase (AMCase) [68].…”

“…In order to study the interaction of RAS modulators or anti-inflammatory compounds with SARS-CoV-2 Spike RBD-Fc protein, cells were pretreated or not pretreated with the AT1 receptor blocker Candesartan (1 μM; 4791, Tocris) or the ACE inhibitor Captopril (50 ng/ml; C4042, Sigma) for 24 h. Then, 1 µg/ml of SARS-CoV-2 Spike RBD-Fc protein (40592-V02H, Sino Biological) was added to the cells for 3 h at 37°C. Doses were stablished based on preliminary experiments and previous in vitro studies [ 32 , 33 ].…”

Exosomes from COVID-19 Patients Carry Tenascin-C and Fibrinogen-β in Triggering Inflammatory Signals in Cells of Distant Organ

“…The presence of SARS-CoV-2 RNA in blood was reported in very low number of postmortem samples [ 2 ] and these observations are debatable [ 3 ]. The pathophysiology of extrapulmonary manifestations is not entirely clear, but thought to be due in part to a dysregulated inflammatory response characterized by inhibition of interferon signaling by the virus, T cell lymphodepletion, and the production of pro-inflammatory cytokines, particularly interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) [ 4 , 5 ]. Extrapulmonary manifestations of SARS-CoV-2-associated disease have been documented in numerous organ systems including, but not limited to, cardiac, neurologic, hemostatic, kidney, and liver.…”