Figure 3

Pantethine preentry treatment reduced SARS-CoV-2 infection in Vero E6 cell cultures. Preentry treatment with pantethine reduced the infection of Vero E6 cells by SARS-CoV-2 (MOI 0.05) significantly and in a dose-dependent manner. (A) Seventy-two hours post-infection, cells were collected and stained to determine viability and infection rates through the detection of the viral spike (S) protein in cells by flow cytometry analysis. The presented data, with the percentage of infection in each plot, are representative of 1 of 3 independent experiments that yielded similar results. (B) Data represent the percentage of infection observed with the cytometry-analysis experiments and are shown as the mean + SEM of results obtained from 3 independent experiments with 3 independent points per condition. (C) Seventy-two hours post-infection, cells were lysed with RIPA buffer, and western blot analyses were performed to detect the expression of the viral spike (S), full-length S1 domain, and nucleocapsid (N) proteins. GAPDH was used as a loading control. The numbers below the blots represent relative expression levels. For each viral protein, the levels of the uninfected cells were set at 1. (D) Virus yields in the supernatant of infected cells were quantified by qRT-PCR for the viral N gene. Calculated Ct values were converted to fold-reduction of samples compared to the noninfected cells using the ΔΔCt method (fold change in viral RNA = 2^−ΔΔCt). In (B) and (D), the results represent the mean + SEM. (E) An inhibitory dose–response curve based on viral N gene expression in supernatants was used to determine the IC50 using GraphPad Prism 8 software. The results represent the mean ± SEM. In all experiments, the results were obtained from 3 independent experiments with 3 independent points per condition. **p < 0.01, ***p < 0.001 and ****p < 0.0001 compared to the control group (infected untreated cells) by one-way ANOVA followed by Dunnett's post hoc test.