PM2.5 Exposure Facilitates SARS-CoV-2 Infection through ACE2/TMPRSS2 Regulation and Suppression of Anti-Viral Response

Background: Epidemiological studies suggest an interaction between air pollution including particulate matter <2.5 μm (PM2.5) and Coronavirus disease 2019 (COVID-19) mortality and morbidity; however, the underlying mechanisms are not clear. The aim of our study was to investigate effects of PM2.5 on viability, epithelial integrity, and cellular entry of SARS-CoV-2 into airway epithelial cells, and the mechanisms involved. Methods: We exposed Calu-3 airway epithelial cell cultures to PM2.5 (10, 50, and100 μm/ml) and SARS-CoV-2 (MOI 0.01) for 24 h. The viability of Calu-3 cells and epithelial barrier integrity were determined using MTT assay and immunofluorescence staining for Zonula Occludens-1, respectively. mRNA expression for inflammatory and inflammasomal genes including interleukin (IL)-8, nuclear factor (NF)-kB p65 (RelA), JNK, c-JUN, Caspase-1 and NLRP3, and viral entry-related genes such as angiotensin converting enzyme (ACE)2 and transmembrane protease, serine (TMPRSS)2 were analyzed by qRT-PCR. Intracellular viral spike protein intensity and RNA-dependent RNA polymerase (RdRp) expression were determined using immunofluorescence staining and qRT-PCR, respectively. Inflammatory cytokine release was analyzed by ELISA. Results: Higher concentrations of 100μg/ml PM2.5 decreased Calu-3 cell viability (p=0.02) and deteriorated epithelial barrier integrity, and 50 μg/ml of PM2.5 (p<0.01) induced mRNA expression for ACE2 and TMPRSS2. Although PM2.5 alone did not alter the expression of mRNA for JNK and RelA, a combination of SARS-CoV-2 and 50-100μg/ml PM2.5 led to a significant increase in mRNA for both genes (p<0.05 and p<0.01, respectively). Moreover, our results indicated an increasing expression of IL-6 and GM-CSF by PM2.5 and PM2.5 + SARS-CoV-2 exposure, whereas IL-8 was induced by SARS-CoV-2. Conclusion: Our findings demonstrated that PM2.5 impaired epithelial integrity and viability, whereas increased the mRNA expression for ACE2 and TMPRSS2, and induced inflammatory changes in Calu-3 cells incubated with SARS-CoV-2. These findings suggest that PM2.5 can induce airway epithelial cell entry of SARS-CoV-2, and that both PM2.5 and SARS-CoV-2 can interact in causing the inflammatory response of the host cell.