The spike protein of the causative COVID-19 virus induces heme oxygenase-1: Pathophysiologic implications

ABSTRACT

Background: Acute kidney injury (AKI) is both a consequence and determinant of outcomes in COVID-19. The kidney is one of the major organs infected by the causative virus SARS-CoV-2. The spike protein of SARS-CoV-2 is required for viral entry into cells and is present in the urine of patients with COVID-19 and AKI. The present study examined cellular effects that result from transfecting the spike protein of SARS-CoV-2 in HEK293 kidney cells. Methods: HEK293-ACE2+ cells stably overexpressing ACE2 were used. Codon optimized pcDNA encoding SARS-CoV-2 spike (7788bp) or empty vector (4033bp) plasmid was transfected using Lipofectamine LTX. For studies examining the effect of quercetin (an inducer of heme oxygenase-1, HO-1), full media containing quercetin or vehicle was added at 4-6 hours post transfection. mRNA and protein expression was assessed by quantitative real-time RT-PCR and western blot respectively. Syncytium formation was assessed by acquiring phase contrast images using Olympus CK40 microscope and the area covered by syncytia was measured using ImageJ software. Results: HEK293-ACE2+ cells expressed SARS-CoV-2 spike protein upon spike transfection. Such expression led to syncytia formation, the sloughing of sheets of cells, and focal denudation of the cell monolayer. Spike protein expression upregulated potentially nephrotoxic genes such as TNF-α, MCP-1, and ICAM1. Spike protein expression also upregulated potentially cytoprotective genes such as HO-1, as demonstrated by HO-1 mRNA and protein expression and relevant signaling pathways (p-Akt, p-STAT3, and p-p38) involved in inducing the HO-1 gene. Quercetin, a naturally occurring compound that induces HO-1, markedly reduced syncytia formation and spike protein expression. Conclusions: These findings introduce a clinically relevant, spike protein-induced, in vitro model for the study of AKI in COVID-19. The major conclusions of the study are 1) Spike protein expression in kidney cells provides a useful and timely model for the study of maladaptive and adaptive responses in these cells relevant to AKI observed in COVID-19;2) spike protein expression in kidney cells upregulates HO-1;and 3) quercetin, an inducer of HO-1, may provide a clinically relevant/feasible protective strategy in AKI occurring in the setting of COVID-19.