Elevated circulating monocytes and monocyte activation in COVID-19 convalescent individuals

Abstract

Background: Monocytes and macrophages play a pivotal role in inflammation during acute SARS-CoV-2 infection. However, their contribution to the development of post-acute sequelae of SARS-CoV-2 infection (PASC) are not fully elucidated.

Methods: A cross-sectional study was conducted comparing plasma cytokine and monocyte levels among three groups: participants with pulmonary PASC (PPASC) with a reduced predicted diffusing capacity for carbon monoxide [DLCOc, <80%; (PG)]; fully recovered from SARS-CoV-2 with no residual symptoms (recovered group, RG); and negative for SARS-CoV-2 (negative group, NG). The expressions of cytokines were measured in plasma of study cohort by Luminex assay. The percentages and numbers of monocyte subsets (classical, intermediate, and non-classical monocytes) and monocyte activation (defined by CD169 expression) were analyzed using flow cytometry analysis of peripheral blood mononuclear cells.

Results: Plasma IL-1Ra levels were elevated but FGF levels were reduced in PG compared to NG. Circulating monocytes and three subsets were significantly higher in PG and RG compared to NG. PG and RG exhibited higher levels of CD169⁺ monocyte counts and higher CD169 expression was detected in intermediate and non-classical monocytes from RG and PG than that found in NG. Further correlation analysis with CD169⁺ monocyte subsets revealed that CD169⁺ intermediate monocytes negatively correlated with DLCOc%, and CD169⁺ non-classical monocytes positively correlated with IL-1α, IL-1β, MIP-1α, Eotaxin, and IFN-γ.

Conclusion: This study present evidence that COVID convalescents exhibit monocyte alteration beyond the acute COVID-19 infection period even in convalescents with no residual symptoms. Further, the results suggest that monocyte alteration and increased activated monocyte subsets may impact pulmonary function in COVID-19 convalescents. This observation will aid in understanding the immunopathologic feature of pulmonary PASC development, resolution, and subsequent therapeutic interventions.

Keywords: CD169; Long-COVID; SARS-CoV-2; monocytes; post-acute sequalae of SARS-CoV-2 infection; pulmonary sequelae.

Figure 1

Plasma cytokine levels among groups. (A) IL-1Ra, (B) PDGF-AA, (C) PDGF-AB/BB, (D) TGF-α, and (E) FGF levels were measured in plasma collected from NG, RG, and PG. The data are represented as the box and scatter plots with each circle representing a single individual. P values were calculated using the Mann–Whitney U-test. *p < 0.05, **p < 0.01, ns, non-significant.

Figure 2

Comparison of circulating monocyte levels among the groups. Representative flow cytometry gating strategy for identification of monocytes and monocyte subsets in PBMC from NG, RG, and PG groups. Lymphocytes and monocytes were selected using CD45⁺ followed by gating for CD11b⁺ cells. (A1) non-classical (CD14^lo/CD16⁺), (A2) intermediate (CD14⁺/CD16⁺, (A3) Classical (CD14⁺/CD16^-), and (B) Total monocyte percentages as a proportion of total identified CD45⁺ cells in NG, RG, and PG groups. (C) Total monocyte counts in NG, RG, and PG groups. Mann-Whitney-U Test *p < 0.05, **p < 0.01, ***p < 0.001, ns, non-significant.

Figure 3

Comparison of circulating monocyte subsets among the groups. (A) Classical monocyte percentage, (B) Intermediate monocyte percentage, (C) Non-Classical monocyte percentage, as a proportion of total identified CD45⁺ cells in NG, RG, and PG groups. (D) Classical monocyte counts, (E) Intermediate monocyte counts, and (F) Non-Classical monocyte in NG, RG, and PG groups. Mann-Whitney-U Test *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, ns, non-significant.

Figure 4

Characterization of circulating CD169⁺ monocytes in NG, RG, and PG groups. Representative histogram and dot plot of CD169⁺/CD14⁺ monocytes in NG, RG, and PG groups. Percentage of total CD169⁺ monocytes from total CD14⁺ monocytes is shown above the gate (B). Total CD169⁺ monocyte percentage from CD45⁺ cells, (C) CD169⁺ monocyte, (D) MFI of CD169 on monocyte subsets in NG, RG, and PG. The percentage of CD169⁺ cells identified in (E) classical monocytes, (F) intermediate monocytes, and (G) non-classical monocytes within NG, RG, and PG groups. Mann-Whitney-U Test *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, ns, non-significant.

Figure 5

Spearman correlation between DLCOc% and (A) percentage of CD169⁺ total monocytes, (B) percentage of CD169⁺ intermediate monocytes, and (C) CD169⁺ intermediate monocyte count in PG.

Figure 6

CD169⁺ non-classical monocytes were associated with IL-1α, IL-1β, MIP-1α, Eotaxin, and IFNγ in PG. Spearman correlation between the percentage of non-classical CD169⁺ monocytes and (A) IL-1α and (B) IL-1β. Spearman correlation between CD169⁺ non-classical monocyte count and (C) MIP-1α, (D) Eotaxin, (E) IL-1α, and (F) IFNγ.