Alteration of serum markers in COVID‐19 and implications on mortality

Dear editor, Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has emerged as a global pandemic since its initial outbreak in Wuhan, China.1 Among the many unanswered questions for COVID-19, how to reduce mortality and improve survival of patients is the most critical. Evidence indicates that increased cytokine levels (also known as cytokine storm) might be a major contributor of disease deterioration in COVID-19 leading to death.2–4 Previous reports focused on cytokine storm in COVID-19 have been limited by shorter followups, small sample sizes, and evaluation of subgroups of patients.5–8 Longitudinal profile of cytokine variations in large cohort has not been comprehensively evaluated. Here, we present an analysis of laboratory indices and cytokines, along with risk factors and mortality in patients with COVID-19 using a closed, multicenter retrospective cohort study. Overall, 2044 COVID-19 patients, who hospitalized in theOptical Valley Campus and Sino-FrenchNew City Campus of Tongji Hospital inWuhan, China, between January 27 and March 21 and had definite outcome (discharge or death), were included in our analysis (Figure S1). Overall, 235 patients died during hospitalization, while 1809 patients were discharged from the hospital (Table S1). The median (IQR) age of all patients was 62.0 (IQR 51.070.0) years, and 48.92%weremen. Over half of the patients (1175, 57.63%) had at least one comorbidity. Patients who died were significantly older (median (IQR) age, 70 [6378] vs 61 [49-69], P < .0001), more likely to be male (156 [66.38%] vs 844 [46.66%], P < .001) and to have comorbidities (189 [81.12%] vs 986 [54.60%], P < .001). Cytokine profiles of patients are shown in Table S2. Over half of COVID-19 patients (1110, 57.33%) had increased C reactive protein (CRP) on admission. Elevated ferritin

and tumor necrosis factor-α (TNF-α) occurred in 651 (56.17%) and 762 (46.15%) patients, respectively. Elevation of cytokines such as IL-2R and IL-6 occurred in 589 (35.61%) and 547 (32.89%) patients, respectively. Patients who died had significantly higher median levels of CRP (103.  Table S3. To depict the dynamic course of COVID-19 and further explore the risk factors associated with poor prognosis, cytokines and other significant indices were tracked over 6 weeks (shown in Figures S2 and S3). The mortality reached its peak approximately around the third week of illness from the onset. Similarly, elevations of D-dimer and cTnI were also observed around 3 weeks from the illness onset among patients with higher mortality. Lastly, in week 5-6, elevated levels of cytokines (including IL-2R, IL-6, IL-8, and IL-10, Ferritin, and TNF-α) were seen among non-survivors compared with patients who recovered. Other laboratory indices, such as LDH, procalcitonin (PCT), and NT-proBNP, also peaked in the corresponding period.
To further understand the role of cytokine storm and other risk factors in COVID-19-related mortality, univariable logistic regression was performed as summarized in Table 1. Older patients were associated with increased odds of death than the younger. The risk of death increased proportionately with the number of comorbidities. The presence of severe respiratory symptoms and unstable vital signs on admission also predicted poor outcomes.
Surprisingly, we found that some risk factors of mortality disproportionately affected males compared to females. Hypertension (OR 2.88, 95% CI 2.02 to 4.11, P < .001) and coronary heart disease (CHD) (2.96, 1.88 to 4.67, P < .001) increased the odds of death in males but had no

F I G U R E 1 Risk Management during the Clinical Course of COVID-19
. Different strategies of risk management in different periods of COVID-19. Leukocytosis was defined as white blood cells count greater than 10 × 10 9 /L. Lymphocytopenia was defined as lymphocyte count less than 0.8 × 10 9 /L. Risk factors for male: age ≥ 50 years, with hypertension, D-dimer > 1 μg/mL, PCT ≥ 0.25 ng/mL, CRP > 10 mg/L, leukocytosis, lymphocytopenia, and IL-6 ≥ 14 pg/mL. Risk factors for female: IL-8 ≥ 62 pg/mL, IL-2R > 710 U/mL, cTnI > 15.6 pg/mL, leukocytosis, lymphocytopenia, and IL-6 ≥ 14 pg/mL. a Time from illness onset adverse influence on females ( Figures S4 and S5). However, women with cancer and COPD had a higher risk of death compared to men. Females were also found to have increased odds of death in the cases of cytokine storm, cardiac injury, and coagulopathy compared to their male counterparts.
In the multivariable logistic regression model, we analyzed the factors with significant impact on mortality separately for males and females. Overall, IL-6 ≥ 14 pg/mL, leukocytosis, and lymphocytopenia were independent risk factors of death for both sexes. For male patients, advanced age (≥50 years), hypertension, the elevated PCT, CRP, and D-dimer were associated with increased odds of mortality. Meanwhile, IL-2R, IL-8, and cTnI were independent risks for higher mortality in females.
To further investigate the influence of risk factors, we plotted Kaplan-Meier curves to study the prognoses of patients based on the numbers of independent risk factors they had ( Figure S4C,D). Males (females) who had 0-3 (0-2), 4-6 (3), and 7-8 (4-6) factors were regarded as lowrisk, moderate-risk, and high-risk patients, respectively, and had diverse prognoses. 67.00% (75.93%) of the high-risk male (female) patients died while only 0.82% (0.77%) of the low-risk ones had fatal outcome. A summary of these findings is shown in Figure 1.
In conclusions, IL-6 ≥ 14 pg/mL, leukocytosis, and lymphocytopenia were found to be risk factors for mortality in both sexes. Advanced age, presence of hypertension, elevated D-dimer, CRP, and PCT were independent risk factors of death only in males, while elevated IL-2R, IL-8, and cTnI increased the risk of mortality in females. Increase in D-dimer and cTnI were observed in the second and third weeks of illness onset while multiple cytokines were found to be increased in the fifth and sixth weeks among those with high mortality. Cytokine storm was a major concern throughout the clinical course, especially in later stages of COVID-19 and among females. Whether early intervention with potential anti-inflammatory or anti-cytokine agents can improve the prognosis in COVID-19 remains to be seen. Risk stratification based on cytokine profile and other risk factors might be considerable in the management of COVID-19.