Liquid biomarkers of macrophage dysregulation and circulating spike protein illustrate the biological heterogeneity in patients with post-acute sequelae of COVID-19

Abstract

Post-acute sequelae of COVID-19 (PASC) are long-term consequences of SARS-CoV-2 infection that can substantially impair the quality of life. Underlying mechanisms ranging from persistent viruses to innate and adaptive immune dysregulation have been discussed. Here, we profiled the plasma of 181 individuals from the cohort study for digital health research in Germany (DigiHero), including individuals after mild to moderate COVID-19 with or without PASC and uninfected controls. We focused on soluble factors related to monocyte/macrophage biology and on circulating SARS-CoV-2 spike (S1) protein as a potential biomarker for persistent viral reservoirs. At a median time of 8 months after infection, we found pronounced dysregulation in almost all tested soluble factors, including both pro-inflammatory and pro-fibrotic cytokines. These immunological perturbations were remarkably independent of ongoing PASC symptoms per se, but further correlation and regression analyses suggested PASC-specific patterns involving CCL2/MCP-1 and IL-8 that either correlated with sCD162, sCD206/MMR, IFN-α2, IL-17A and IL-33, or IL-18 and IL-23. None of the analyzed factors correlated with the detectability or levels of circulating S1, indicating that this represents an independent subset of patients with PASC. These data confirm prior evidence of immune dysregulation and persistence of viral protein in PASC and illustrate its biological heterogeneity that still awaits correlation with clinically defined PASC subtypes.

Keywords: COVID-19; SARS coronavirus; cytokine/chemokine; long COVID; macrophage; monocyte.

Figure 1

Clinical and epidemiological characteristics of the post‐COVID‐19 cohort. (A) Sex distribution in the analyzed cohort comprised individuals with ongoing PASC (n = 91), individuals with prior COVID‐19 who never reported PASC (n = 62), and individuals without prior COVID‐19 (n = 28). (B) Violin plot of median blood sampling time point (continuous line) relative to positive polymerase chain reaction or antigen test for the post‐COVID‐19 groups. Dotted lines separate quartiles. (C) Median age of indicated groups.  PASC, post‐acute sequelae of COVID‐19.

Figure 2

Profiling of plasma monocyte/macrophage‐related soluble factors from individuals with ongoing PASC, without PASC, and without SARS‐COV‐2 infection. Mean plasma cytokine/chemokine/soluble factor levels of individuals with no prior COVID‐19 (n = 28), individuals who never reported PASC postinfection (n = 62), or with ongoing PASC (n = 91). Error bars indicate ±SD. Statistical analysis: Welch's analysis of variance for comparison of all three groups and two‐sided Welch corrected t‐test for comparison of the no prior COVID‐19 versus never PASC, no prior COVID‐19 versus ongoing PASC, and never PASC versus ongoing PASC groups. PASC, post‐acute sequelae of COVID.

Figure 3

Correlation analysis of plasma soluble factors. (A–C) Correlation matrix of all analyzed soluble factors for individuals with ongoing PASC (n = 91) (A), individuals who never reported PASC postinfection (n = 62) (B), or individuals with no prior COVID‐19 (n = 28) (C). PASC, postacute sequelae of COVID.

Figure 4

Association of plasma soluble factors with sampling time point postinfection. (A, B) Linear regression of plasma cytokine levels and sampling time point postinfection in individuals without PASC (n = 62) (A) and with ongoing PASC (n = 91) (B). Both cohorts also comprise cytokine data from individuals with mild/moderate acute COVID‐19 (n = 15). The dotted red lines indicate the mean plasma level determined in individuals without prior COVID‐19 (n = 28). Correlation coefficient R ², Spearman correlation coefficients (r _S), and p values are indicated. PASC, post‐acute sequelae of COVID.

Figure 5

Persistence of circulating S1 protein and correlation with monocyte/macrophage‐related soluble factors and SARS‐CoV‐2‐directed antibodies in unvaccinated individuals with ongoing PASC. (A) Proportion of individuals with detectable levels of circulating S1 (cS1) protein in unvaccinated individuals with prior COVID‐19 who never experienced PASC (n = 17) and with ongoing PASC (n = 22). (B) Mean plasma levels of circulating SARS‐CoV‐2 spike (S1) protein in unvaccinated individuals with ongoing PASC (n = 22), individuals with prior COVID‐19 who never reported PASC (n = 17), and individuals without prior COVID‐19 (n = 2). Error bars indicate ±SD. Statistical analysis: one‐sided Welch corrected t‐test. (C) Correlation matrix of indicated soluble factors with levels of circulating S1 and S1/NCP antibodies in unvaccinated individuals with ongoing PASC. NCP, nucleocapsid protein; PASC, post‐acute sequelae of COVID.