Native High-Density Lipoproteins (HDL) with Higher Paraoxonase Exerts a Potent Antiviral Effect against SARS-CoV-2 (COVID-19), While Glycated HDL Lost the Antiviral Activity

Abstract

Human high-density lipoproteins (HDL) show a broad spectrum of antiviral activity in terms of anti-infection. Although many reports have pointed out a correlation between a lower serum HDL-C and a higher risk of COVID-19 infection and progression, the in vitro antiviral activity of HDL against SARS-CoV-2 has not been reported. HDL functionality, such as antioxidant and anti-infection, can be impaired by oxidation and glycation and a change to pro-inflammatory properties. This study compared the antiviral activity of native HDL with glycated HDL via fructosylation and native low-density lipoproteins (LDL). After 72 h of fructosylation, glycated HDL showed a typical multimerized protein pattern with an elevation of yellowish fluorescence. Glycated HDL showed a smaller particle size with an ambiguous shape and a loss of paraoxonase activity up to 51% compared to native HDL. The phagocytosis of acetylated LDL was accelerated 1.3-fold by glycated HDL than native HDL. Native HDL showed 1.7 times higher cell viability and 3.6 times higher cytopathic effect (CPE) inhibition activity against SARS-CoV-2 than that of glycated HDL under 60 μg/mL (approximately final 2.2 μM) in a Vero E6 cell. Native HDL showed EC₅₀ = 52.1 ± 1.1 μg/mL (approximately final 1.8 μM) for the CPE and CC₅₀ = 79.4 ± 1.5 μg/mL (around 2.8 μM). The selective index (SI) of native HDL was calculated to be 1.52. In conclusion, native HDL shows potent antiviral activity against SARS-CoV-2 without cytotoxicity, while the glycation of HDL impairs its antiviral activity. These results may explain why patients with diabetes mellitus or hypertension are more sensitive to a COVID-19 infection and have a higher risk of mortality.

Keywords: COVID-19; SARS-CoV-2; glycation; high-density lipoproteins (HDL); low-density lipoproteins; paraoxonase.

Figure 1

Characterization of HDL. (A) Electrophoretic patterns of native HDL and glycated HDL (12% SDS-PAGE). Lane M, molecular weight standard (Bio-Rad, low-range). (B) Fluorospectroscopic determination of glycation extent in HDL during 72 h (hrs).

Figure 2

Observation of the HDL particle and antioxidant activity. (A) Negatively stained image of HDL from transmitted electron microscopy at a magnification of ×40,000. Scale bar (yellow) indicates 100 nm. (B) Changes in the activity of paraoxonase in HDL with or without glycation. The error bars indicate the SD from three independent experiments with duplicate samples. The molar extinction coefficient of p-nitrophenol was 17,000 M⁻¹ cm⁻¹. ***, p < 0.001.

Figure 3

Anti-atherosclerotic activity with HDL. (A) Cellular uptake of acLDL in the presence of each HDL. Photo a, PBS alone; Photo b, acLDL-alone; Photo c, acLDL + native-HDL; Photo d, acLDL + glycated-HDL. (B) Determination of oxidized species in the cell media as a malondialdehyde standard. Oil-red O-stained cellular areas of 0.8 mm² were quantified using computer-assisted morphometry. Data are shown as the mean ± SD of three independent experiments performed in triplicate.

Figure 4

Comparison of glycated HDL and native HDL. (A) Cell viability of Vero E6 cell after treatment with lipoproteins. MTT cytotoxicity assays were carried out using uninfected Vero E6 cells with increasing concentrations of each lipoprotein. (B) Cytopathic effect (CPE) reduction activity of lipoproteins against SARS-CoV-2.

Figure 5

Vero E6 Cell viability (A) and cytopathic effect (CPE) reduction activity (B) of native HDL. EC₅₀, effective concentration of compound needed to inhibit the CPE to 50% of control value. CC₅₀, the cytotoxic concentration of the compound that reduced cell viability to 50%. SI, selectivity index = EC₅₀/CC₅₀.