Prediction of Post-Acute-Sequelae of COVID-19 by Cargo Protein Biomarkers of Blood Total Extracellular Vesicles in Acute COVID-19

Abstract

Background: SARS-CoV-2 invades mitochondria of infected cells resulting in disordered metabolism, mitophagy, and abnormal levels of mitochondrial proteins in extracellular vesicles. Blood extracellular vesicle SARS-CoV-2 proteins and mitochondrial proteins were quantified in COVID-19 to assess possible roles as biomarkers.

Methods: Total extracellular vesicles were precipitated from blood of age- and gender-matched participants with no infection (n=10), acute COVID-19 (n=16), post-acute sequelae of COVID-19 (PASC or long COVID) (n=30), or post-acute COVID without PASC (n=8) and their extracted proteins quantified by enzyme-linked immunosorbent assays (ELISAs).

Results: Total extracellular vesicle levels of S1 (receptor-binding domain [RBD]) protein were significantly higher in acute infections than in uninfected controls, post-acute infection without PASC, and PASC. Total extracellular vesicle levels of nucleocapsid (N) protein were significantly higher in PASC than in uninfected controls, acute infections, and post-acute infection without PASC. Neither acute levels of S1(RBD) or N proteins predicted progression to PASC. Levels of neither SARS-CoV-2 protein in established PASC correlated with neuropsychiatric manifestations. Significant decreases in total extracellular vesicle levels of the mitochondrial proteins MOTS-c, VDAC-1, and humanin, and elevations of levels of SARM-1 were observed in acutely infected patients who would develop PASC. Significant decreases in total extracellular vesicle levels of MOTS-c and humanin, but not VDAC-1, and elevations of total extracellular vesicle levels of SARM-1 were characteristic of PASC patients with neuropsychiatric manifestations.

Conclusions: Total extracellular vesicle levels of SARS-CoV-2 proteins in COVID-19 indicate intracellular presence of SARS-CoV-2. Abnormal total extracellular vesicles levels of mitochondrial proteins in acute infections predict a high risk of PASC and later in established PASC are indicative of neuropsychiatric manifestations.

Keywords: Long COVID; Mitochondria; Neuropsychiatric effects of COVID-19; SARS-CoV-2 proteins S1 and N.

Figure 1

Plasma total extracellular vesicle (EV) levels of SARS-CoV-2 proteins. (A) S1 (receptor-binding domain) protein and (B) N protein. Each point represents the CD81-normalized value for 1 participant, and the horizontal line in each cluster of points is the mean value for that group. “Uninfected Ctl” are the controls and statistical comparators for “Acute to PASC,” “Acute to resolution,” and “Post-acute w/o PASC” groups. “Post-acute w/o PASC” are the controls and statistical comparators for “PASC w/NP” and “PASC w/o NP groups". P values from unpaired t tests are *P < .01 and **P < .001.

Figure 2

Plasma total extracellular vesicle (EV) levels of mitochondrial proteins. (A) MOTS-c, (B) SARM-1, (C) VDAC-1, (D) humanin, (E) LETM-1, and (F) CI-6. Each point represents the CD81-normalized value for 1 participant, and the horizontal line in each cluster of points is the mean value for that group. “Uninfected Ctl” are the controls and statistical comparators for “Acute to PASC,” “Acute to resolution,” and “Post-acute w/o PASC” groups. “Post-acute w/o PASC” are the controls and statistical comparators for “PASC w/NP” and “PASC w/o NP groups”. The P values from unpaired t tests are *P < .01 and **P < .001.