Immune-Mediated Inflammatory Responses of Alveolar Epithelial Cells: Implications for COVID-19 Lung Pathology

Abstract

Background. Clinical and experimental evidence point to a dysregulated immune response caused by SARS-CoV-2 as the primary mechanism of lung disease in COVID-19. However, the pathogenic mechanisms underlying COVID-19-associated ARDS (Acute Respiratory Distress Syndrome) remain incompletely understood. This study aims to explore the inflammatory responses of alveolar epithelial cells to either the spike S1 protein or to a mixture of cytokines secreted by S1-activated macrophages. Methods and Results. The exposure of alveolar A549 cells to supernatants from spike-activated macrophages caused a further release of inflammatory mediators, with IL-8 reaching massive concentrations. The investigation of the molecular pathways indicated that NF-kB is involved in the transcription of IP-10 and RANTES, while STATs drive the expression of all the cytokines/chemokines tested, with the exception of IL-8 which is regulated by AP-1. Cytokines/chemokines produced by spike-activated macrophages are also likely responsible for the observed dysfunction of barrier integrity in Human Alveolar Epithelial Lentivirus-immortalized cells (hAELVi), as demonstrated by an increased permeability of the monolayers to mannitol, a marked decrease of TEER and a disorganization of claudin-7 distribution. Conclusion. Upon exposure to supernatants from S1-activated macrophages, A549 cells act both as targets and sources of cytokines/chemokines, suggesting that alveolar epithelium along with activated macrophages may orchestrate lung inflammation and contribute to alveolar injury, a hallmark of ARDS.

The JAK1/2 Inhibitor Baricitinib Mitigates the Spike-Induced Inflammatory Response of Immune and Endothelial Cells In Vitro

“…Similar results have been shown in vivo by Bronte et al, who reported a tendency of IL-8 plasma levels to increase after 7 d of treatment with baricitinib, despite an evident drop of IL-6 during the same time [ 29 ]. IL-8 expression in coronavirus infected cells has been reported under AP-1 control [ 47 ], and, similarly, we found the same transcription factor involved in IL-8 synthesis in epithelial A549 cells [ 48 ]. Given the primary role of this chemokine as a potent attractant for neutrophils [ 49 , 50 ], the inefficacy of baricitinib on IL-8 release could explain why the transendothelial migration of neutrophils in vitro is stimulated by the CM_S1 medium and unaffected by baricitinib.…”

Hesperetin from Root Extract of Clerodendrum petasites S. Moore Inhibits SARS-CoV-2 Spike Protein S1 Subunit-Induced NLRP3 Inflammasome in A549 Lung Cells via Modulation of the Akt/MAPK/AP-1 Pathway

“…In addition to pro-inflammatory cytokine release, previous studies have reported that the NLRP3 inflammasome pathway activated by Spike S1 led to an increase in pro-inflammatory cytokine gene expressions, including NLRP3, IL-1β and IL-18 [ 34 , 35 , 36 , 37 ]. Thus, the effects of the C. petasites extracts, CpEE and CpEA, as well as hesperetin on NLRP3 , IL-1β , and IL-18 mRNA expressions in Spike S1-induced A549 cells, were also determined using RT-qPCR.…”

“…This could eventually lead to the chronic or sub-chronic systemic inflammation that was observed in COVID-19 patients during the post-infection phase [ 3 , 12 ]. With regard to our in vitro investigation of the SP-induced inflammatory mechanism, as well as the therapeutic potential of the C. petasites extract and its active compounds, we selected A549 cells as our cell culture model because ACE2-expressed airway epithelial cells were used in the experiments of previous studies that focused on SARS-CoV-2 infection and inflammation [ 35 , 36 , 37 ]. Accordingly, these aforementioned studies used the same A549 cells to represent the lower respiratory system or the alveolar epithelial cells that were induced by the spike glycoprotein.…”

“…Nowadays, many studies have confirmed the role of the spike glycoprotein of SARS-CoV-2 on the inflammatory induction during COVID-19 infections. Briefly, the spike glycoprotein S1 was found to trigger an inflammatory response in human PBMC, monocyte, and lung epithelial cell lines [ 34 , 35 , 36 , 40 ]. Additionally, in our recent study, 100 ng/mL of the spike glycoprotein S1 was able to induce NLRP3 cytokine releases, including IL-6, IL-1β, and IL-18, in both gene and protein levels for A549 lung epithelial cells and PMA-treated THP-1 macrophages [ 37 ].…”

Cyanidin-3-O-glucoside and Peonidin-3-O-glucoside-Rich Fraction of Black Rice Germ and Bran Suppresses Inflammatory Responses from SARS-CoV-2 Spike Glycoprotein S1-Induction In Vitro in A549 Lung Cells and THP-1 Macrophages via Inhibition of the NLRP3 Inflammasome Pathway

“…To validate the inflammatory response conditions in both A549 lung cells and PMAtreated THP-1 macrophages, we first determined the optimal time required for an inflammatory response upon SP induction. A concentration of SP at 100 ng/mL was chosen from the review literature of previous studies that were investigated on inflammatory responses in lung epithelial cells [7,36], monocytes, or macrophage cells [3,5]. This concentration was then confirmed in our own laboratory.…”