High Circulating Levels of the Homeostatic Chemokines CCL19 and CCL21 Predict Mortality and Disease Severity in COVID-19

Abstract

Background: Immune dysregulation is a major factor in the development of severe coronavirus disease 2019 (COVID-19). The homeostatic chemokines CCL19 and CCL21 have been implicated as mediators of tissue inflammation, but data on their regulation in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is limited. We thus investigated the levels of these chemokines in COVID-19 patients.

Methods: Serial blood samples were obtained from patients hospitalized with COVID-19 (n = 414). Circulating CCL19 and CCL21 levels during hospitalization and 3-month follow-up were analyzed. In vitro assays and analysis of RNAseq data from public repositories were performed to further explore possible regulatory mechanisms.

Results: A consistent increase in circulating levels of CCL19 and CCL21 was observed, with high levels correlating with disease severity measures, including respiratory failure, need for intensive care, and 60-day all-cause mortality. High levels of CCL21 at admission were associated with persisting impairment of pulmonary function at the 3-month follow-up.

Conclusions: Our findings highlight CCL19 and CCL21 as markers of immune dysregulation in COVID-19. This may reflect aberrant regulation triggered by tissue inflammation, as observed in other chronic inflammatory and autoimmune conditions. Determination of the source and regulation of these chemokines and their effects on lung tissue is warranted to further clarify their role in COVID-19.

Clinical trials registration: NCT04321616 and NCT04381819.

Keywords: SARS-CoV-2; chemokine; predictive markers; respiratory distress syndrome; viral infection.

Figure 1.

Flow chart showing the study population, with clinical data and blood samples collected from 2 cohort studies. Further details of the 2 trials are provided in the “Methods” section. Blood samples were collected at 3 time points during hospitalization, and at outpatient follow-up after 3 months. A subset of patients from cohort 1 also underwent pulmonary function assessment and chest CT imaging at follow-up. Abbreviations: CT, computed tomography; Dlco, diffusing capacity of the lungs for carbon monoxide; PCR, polymerase chain reaction; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.

Figure 2.

Intrahospital temporal profile of CCL19 and CCL21 in patients hospitalized with COVID-19 (n = 414) according to (A) respiratory failure (n = 125) or (B) ICU admission (n = 110) during the first 10 days after inclusion. Data is shown as estimated marginal means and 95% CI. The P values reflect the group (outcome) effect from the linear mixed models with subject as random effect, and time and respiratory failure or ICU admission as fixed effects (also as interaction) in addition to age, sex, estimated glomerular filtration rate, and treatment modalities. Shaded areas show reference value range from healthy controls. * P < .05, ** P < .01, *** P < .001 between groups. Abbreviations: BL, baseline; ICU, intensive care unit.

Figure 3.

CCL19 and CCL21 and 60-day mortality in patients hospitalized with COVID-19 (n = 414). A, Kaplan-Meier analysis 60-day mortality (n = 37) according to tertiles (T) of CCL19 (T1 ≤ 1.27 ng/mL, T2 1.28–2.09 ng/mL, T3 > 2.10 ng/mL) and CCL21 (T1 ≤ 1.41 ng/mL, T2 1.42–2.44 ng/mL, T3 > 2.45 ng/mL). B, Temporal profile of CCL19 and CCL21 during the first 10 days after inclusion according to 60-day mortality. Data in B is shown as estimated marginal means and 95% CI. The P values reflect the group (outcome) effect from the linear mixed models with subject as random effect, and time and mortality as fixed effects (also as interaction) in addition to age, sex, estimated glomerular filtration rate, and treatment modalities. Shaded areas show reference value range from healthy controls. * P < .05, ** P < .01, *** P < .001 between groups. Abbreviation: BL, baseline.

Figure 4.

Intrahospital temporal profile of CCL19 and CCL21 according to (A) treatment with hydroxychloroquine (n = 43) and remdesivir (n = 38) as compared with their respective SoC (n = 81) in cohort 1 (NOR Solidarity trial); (B) dexamethasone treatment; (C) COVID-19 wave; and (D) Dlco below or above LLN at 3-month follow-up. Data is shown as estimated marginal means and 95% CI. The P value indicates group effect, and the bold P value indicates the interaction term between time and group from the linear mixed models with subject as random effect, and time and mortality as fixed effects (also as interaction) in addition to age, sex, and estimated glomerular filtration rate. Shaded areas show reference value range from healthy controls. *P < .01, ***P < .001 between groups; †P < .05, ††P < .01 versus wave 3. Abbreviations: Dlco, diffusing capacity of the lungs for carbon monoxide; LLN, lower limit of normal; HCQ, hydroxychloroquine; REM, remdesivir; SoC, standard of care.

Figure 5.

In vitro secretion of homeostatic chemokines in SARS-CoV-2–exposed monocyte-derived dendritic cells. Quantitation of secreted CCL21 (A) and CCL19 (B) in cultures of monocyte-derived dendritic cells after exposure to inactivated SARS-CoV-2 viral particles (0.001 or 0.01 multiplicity of infection [MOI]) for 6 and 24 hours. Results are shown as mean ± SD (n = 3 per treatment condition). *P < .05, independent samples t test.

Figure 6.

Regulation of CCR7, CCR10, CCL19, and CCL21 in public RNAseq analysis data of tissues from COVID-19 patients. A, Differences in CCL19 and CCL21 mRNA expression in lung tissue from COVID-19 patients (n = 19) and controls (n = 3). Source data GSE163529. B, mRNA expression in relation to virus load (VL) in COVID-19 patients (GSE150316, n = 15). C, mRNA expression of CCR7 and CCR10 in peripheral blood mononuclear cells isolated from COVID-19 patients (n = 16) grouped by clinical disease severity (moderate, severe disease, and requiring ICU treatment) and age-/sex-matched healthy controls. Source dataset GSE152418. D, Whole blood leukocytes isolated from patients with COVID-19 (COV19), seasonal coronavirus infection (COV; n = 19), influenza (Inf; n = 17), bacterial pneumonia (Bact; n = 20) and matched healthy controls (n = 19). Source dataset GSE161731. Normalized gene expression quantified as transcripts per million (TPM). A and B, P values are from the group and group*tissue location effects from the mixed model analysis (see description of statistics in Supplementary material). C and D, P values are from the Kruskal-Wallis test with asterisks reflecting the results of the post hoc test. *P < .05, **P < .01, ***P < .001; #P < .01 versus other groups.