“Lung Time No See”: SARS‐Cov‐2 Spike Protein Changes Genetic Expression in Human Primary Bronchial Epithelial Cells After Recovery

Evans, Nicholas; Nichols, Jacob; Pruitt, Kevin; Almodovar, Sharilyn

BACKGROUND: Current reports show that people infected with SARS‐CoV‐2 do not recover completely, and even asymptomatic COVID‐19 patients may experience slight changes in their overall health, which is the basis of the new area of study termed “Long‐Haul COVID”. Our hypothesis is that even in the asymptomatic infection, exposures to the viral spike protein are enough to induce long‐lasting changes in baseline genetic expression. OBJECTIVE: This study sought to survey what type of cell biological processes would be affected in human primary bronchial epithelial cells (HBECs) post‐exposure to spike protein and whether they would persist post‐recovery. METHODS: Herein, we advanced an Air Liquid Interface (ALI) cell culture technique to simulate the physiological conditions in the lung airway in vitro. Briefly, HBECs were grown and differentiated, before treatment with either a low (50 ng/mL) or high (5 ug/mL) concentration of recombinant SARS‐CoV‐2 spike protein for 4 hours. After a 48‐hour recovery, cells were processed for RNA extractions and qPCR to screen genes using Qiagen RT(2) Profiler PCR Arrays; data were analyzed in GeneGlobe. RESULTS: We used the (2^ (‐Delta Delta CT)) method to analyze the gene expression data, using both ACTB and RPLP0 as reference genes for normalization. Our survey included genes related to oxidative stress, hypoxia, osmotic stress, cell death, inflammatory response, DNA damage and unfolded protein response. We found that the genes CCL2, IL1A, IL1B, and MMP9 showed fold changes greater than 2.00 in the low and high concentration treatments after recovery. CONCLUSIONS: Our preliminary results suggest that the SARS‐CoV‐2 spike protein is enough to change the baseline protein expression in primary HBECs. After recovery, genes related to immune response retained changes in gene expression, and these may indicate relevant long‐term effects in asymptomatic patients. Additionally, the interplay between immune response and other pathways after SARS‐CoV‐2 spike protein exposure should be investigated in the future.