Comparison of C-reactive protein with distinct hyperinflammatory biomarkers in association with COVID-19 severity, mortality and SARS-CoV-2 variants

Abstract

C-reactive protein (CRP) has been one of the most investigated inflammatory-biomarkers during the ongoing COVID-19 pandemics caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). The severe outcome among patients with SARS-CoV-2 infection is closely related to the cytokine storm and the hyperinflammation responsible for the acute respiratory distress syndrome and multiple organ failure. It still remains a challenge to determine which of the hyperinflammatory biomarkers and cytokines are the best predictors for disease severity and mortality in COVID-19 patients. Therefore, we evaluated and compared the outcome prediction efficiencies between CRP, the recently reported inflammatory modulators (suPAR, sTREM-1, HGF), and the classical biomarkers (MCP-1, IL-1β, IL-6, NLR, PLR, ESR, ferritin, fibrinogen, and LDH) in patients confirmed with SARS-CoV-2 infection at hospital admission. Notably, patients with severe disease had higher serum levels of CRP, suPAR, sTREM-1, HGF and classical biomarkers compared to the mild and moderate cases. Our data also identified CRP, among all investigated analytes, to best discriminate between severe and non-severe forms of disease, while LDH, sTREM-1 and HGF proved to be excellent mortality predictors in COVID-19 patients. Importantly, suPAR emerged as a key molecule in characterizing the Delta variant infections.

Keywords: COVID-19; CRP; HGF; biomarkers; disease severity; mortality; s-TREM-1; suPAR.

Figure 1

Serum profile of CRP and pro-inflammatory cytokines in mild, moderate and severe COVID-19 disease. (A) Patients’ discharge status (no disease, ameliorated symptoms or deceased) for each category of COVID-19 disease: mild, moderate, severe (***p < 0.001; chi-squared test). Serum levels of (B) CRP, (C) suPAR, (D) sTREM-1, (E) HGF, (F) MCP-1, (G) IL-1β, (H) IL-6 for each category of COVID-19 disease: mild, moderate or severe. The gray lines represent the mean ± SEM (****p < 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05, ns – not significant; (A) chi-squared test; (B-H) Kruskal-Wallis with Dunn’s Multiple Comparison test).

Figure 2

Serum profile of common pro-inflammatory biomarkers in mild, moderate and severe COVID-19 disease. Serum levels of (A) NLR (neutrophil-lymphocyte ratio), (B) PLR (platelet-lymphocyte ratio), (C) ESR (erythrocyte sedimentation rate), (D) fibrinogen, (E) ferritin, (F) LDH for each category of COVID-19 disease: mild, moderate or severe. The gray lines represent the mean ± SEM (****p < 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05, ns – not significant; Kruskal-Wallis with Dunn’s Multiple Comparison test).

Figure 3

Correlation of pro-inflammatory biomarkers with age in mild, moderate and severe SARS-CoV-2 infections. (A) Heat map of correlation coefficients (R) for each category of COVID-19 disease: mild, moderate, severe (0.2-0.39: weak; 0.4-0.59: moderate; 0.6-0.8 strong). (B) CRP levels, (C) suPAR levels, (D) sTREM-1 levels, and (E) NLR values in mild, moderate and severe COVID-19 patients stratified by age (younger or older than 60 years). The gray lines represent the mean ± SEM (***p < 0.001, **p < 0.01, *p < 0.05, ns – not significant; Kruskal-Wallis with Dunn’s Multiple Comparison test).

Figure 4

Regression statistics describing the association between CRP and other pro-inflammatory biomarkers in moderate and severe cases of SARS-CoV-2 infection. (A) Heat map of correlation coefficients (R) for each category of COVID-19 disease: moderate (left-bottom corner) and severe (right-upper corner). Linear regression analysis for (B) CRP and suPAR levels, (C) CRP and sTREM-1 levels, (D) CRP and HGF levels, and (E) sTREM-1 and MCP-1 levels in, moderate and severe COVID-19 patients (****p < 0.0001, **p < 0.01, *p < 0.05, ns – not significant; Spearman test). The blue lines and dots correspond to moderate cases, while the green lines and dots state for severe cases.

Figure 5

Regression statistics describing the association between CRP and classical pro-inflammatory biomarkers in moderate and severe cases of SARS-CoV-2 infection. Linear regression analysis for CRP and (A) ESR, (B) fibrinogen, (C) ferritin, and (D) LDH in moderate and severe COVID-19 patients (****p < 0.0001, ***p < 0.001, **p < 0.01, ns – not significant; Spearman test). The blue lines and dots correspond to moderate cases, while the green lines and dots state for severe cases.

Figure 6

Serum profile of CRP and pro-inflammatory cytokines in Delta and Omicron SARS-CoV-2 infections. Serum levels of (A) suPAR, (B) CRP, (C) sTREM-1, (D) HGF, (E) MCP-1, (F) IL-1β, (G) IL-6 for each category of SARS-CoV-2 infection: Delta or Omicron. The gray lines represent the mean ± SEM (****p < 0.0001, *p < 0.05, ns – not significant; two-tailed Mann-Whitney test). (H) Patients’ discharge status (no disease, ameliorated symptoms or deceased) for each category of SARS-CoV-2 infection: Delta or Omicron (ns – not significant; chi-squared test).

Figure 7

Serum profile of CRP and pro-inflammatory cytokines in severe COVID-19 patients stratified based on survival. Box and whiskers representation of (A) CRP, (B) suPAR, (C) sTREM-1, (D) HGF, (E) MCP-1, (F) IL-1β, (G) IL-6, and (H) LDH serum levels for each category of SARS-CoV-2 severe infection: survived or deceased (**p < 0.01, *p < 0.05, ns – not significant; two-tailed Mann-Whitney test).

Figure 8

Distinct pro-inflammatory biomarkers associate with disease severity, Delta variant infection or increased death rate. (A) Heat map of correlation coefficients of pro-inflammatory biomarkers with disease severity, Delta variant infection or deceased status. ROC curves generated for the (B) association of CRP and other pro-inflammatory biomarkers with severe COVID-19, (C) association of suPAR and other pro-inflammatory biomarkers with Delta variant infection, (D) association of LDH and other inflammatory biomarkers with mortality. The mathematical models from (B–D) are detailed in Supplementary Tables 3–5 , respectively. (E) Kaplan-Meier survival curves for each category of COVID-19 patients: mild, moderate and severe.

Figure 9

Comorbidities associated with COVID-19 severity and mortality. Differences in comorbidity rates (A) between non-severe and severe COVID-19 patients, (B) between survived and deceased COVID-19 patients, and (C) between survived and deceased severe COVID-19 cases (*p < 0.05, ns – not significant; chi-squared test). D = Diseases of the blood involving the immune mechanism (anemia, purpura and other hemorrhagic conditions), E = Endocrine and metabolic disorders, I = Diseases of the circulatory system, K = Diseases of liver and gallbladder.