Association of miR-144 levels in the peripheral blood with COVID-19 severity and mortality

Abstract

Coronavirus disease-2019 (COVID-19) can be asymptomatic or lead to a wide symptom spectrum, including multi-organ damage and death. Here, we explored the potential of microRNAs in delineating patient condition and predicting clinical outcome. Plasma microRNA profiling of hospitalized COVID-19 patients showed that miR-144-3p was dynamically regulated in response to COVID-19. Thus, we further investigated the biomarker potential of miR-144-3p measured at admission in 179 COVID-19 patients and 29 healthy controls recruited in three centers. In hospitalized patients, circulating miR-144-3p levels discriminated between non-critical and critical illness (AUC_miR-144-3p = 0.71; p = 0.0006), acting also as mortality predictor (AUC_miR-144-3p = 0.67; p = 0.004). In non-hospitalized patients, plasma miR-144-3p levels discriminated mild from moderate disease (AUC_miR-144-3p = 0.67; p = 0.03). Uncontrolled release of pro-inflammatory cytokines can lead to clinical deterioration. Thus, we explored the added value of a miR-144/cytokine combined analysis in the assessment of hospitalized COVID-19 patients. A miR-144-3p/Epidermal Growth Factor (EGF) combined score discriminated between non-critical and critical hospitalized patients (AUC_{miR-144-3p/EGF} = 0.81; p < 0.0001); moreover, a miR-144-3p/Interleukin-10 (IL-10) score discriminated survivors from nonsurvivors (AUC_{miR-144-3p/IL-10} = 0.83; p < 0.0001). In conclusion, circulating miR-144-3p, possibly in combination with IL-10 or EGF, emerges as a noninvasive tool for early risk-based stratification and mortality prediction in COVID-19.

Figure 1

Changes in miR-144-3p levels between T₀ and T₁ sampling in critical COVID-19 patients. Patients analyzed were ICU-admitted subjects requiring endotracheal intubation. miR-144-3p levels were measured by qPCR in T₀ (admission) and T₁ (discharge or death) plasma samples of each patient. Values are expressed as log₂ fold change compared to the average values of the first sampling for each group. miR-144-3p expression levels were rescued over time in surviving (a) but not in nonsurviving COVID-19 patients (b). Mann–Whitney test (two groups) was performed for statistical comparison. Survivors n = 12; nonsurvivors n = 7; *p < 0.05.

Figure 2

miR-144 levels are decreased in hospitalized COVID-19 patients and discriminate patients from healthy controls. miR-144-3p (a) and miR-144-5p (b) plasma levels were measured by qPCR. Values are expressed as log₂ fold change compared to controls and shown as dot- plots indicating mean ± SEM. Unpaired t-test (two groups) was used for statistical comparison. The ROC curves show the sensitivity and specificity of miR-144-3p (c) and miR-144-5p (d) to distinguish COVID-19 patients hospitalized at PSD from healthy controls. Controls n = 23; COVID-19 patients n = 97; ****p ≤ 0.0001.

Figure 3

miR-144 levels are decreased in nonsurviving COVID-19 patients and discriminate nonsurviving patients from survivors. miR-144-3p (a) and miR-144-5p (b) plasma levels were measured by qPCR in COVID-19 patients hospitalized at PSD and healthy controls. Values are expressed as log₂ fold change compared to controls and shown as dot-plots indicating mean ± SEM. Both miR-144-3p and miR-144-5p expression levels were lower in nonsurviving patients compared to survivors. ANOVA test, followed by Tukey’s post-hoc test, was performed for statistical comparison. The ROC curves show the sensitivity and specificity of miR-144-3p (c) and miR-144-5p (d) to distinguish surviving from nonsurviving COVID-19 patients. Controls n = 23; survivors n = 42–46; nonsurvivors n = 49–51; **p ≤ 0.01; ***p ≤ 0.001; ****p ≤ 0.0001.

Figure 4

miR-144 levels are decreased in critical COVID-19 patients and discriminate critical from non-critical hospitalized patients. miR-144-3p (a) and miR-144-5p (b) plasma levels were measured by qPCR in hospitalized patients at PSD and healthy controls. Values are expressed as log₂ fold change compared to controls and shown as dot-plots indicating mean ± SEM. Both miR-144-3p and miR-144-5p expression levels decreased in critical compared to non-critical patients. ANOVA test, followed by Tukey’s post-hoc test, was performed for statistical comparison. The ROC curves show the sensitivity and specificity of miR-144-3p (c) and miR-144-5p (d) to distinguish non-critical from critical COVID-19 patients. Controls n = 23; non-critical patients n = 31–32; critical patients n = 60–65; ***p ≤ 0.001; ****p ≤ 0.0001.

Figure 5

miR-144-3p in mild and moderate COVID-19 non-hospitalized patients recruited at LIH. miR-144-3p plasma levels were measured by qPCR. Values are expressed as log₂ fold change compared to mild illness patients and shown as dot- plots indicating mean ± SEM. miR-144-3p expression levels are lower in more ill patients. Mann–Whitney test (two groups) was performed for statistical comparison (a). The ROC curve shows the sensitivity and specificity of miR-144-3p to distinguish mild from moderate COVID-19 patients (b). Mild illness patients n = 59; moderate illness patients n = 17; *p < 0.05.

Figure 6

Discriminating potential of miR-144/cytokine combined analysis in COVID-19 hospitalized patients. The ROC curves show the sensitivity and specificity of miR-144-3p/IL-10 (a) and miR-144-5p/IL-10 (b) scores to distinguish surviving from nonsurviving COVID-19 patients. Survivors n = 42; nonsurvivors n = 20. In panel (c), the ROC curve shows the sensitivity and specificity of miR-144-3p/EGF score to distinguish non-critical from critical COVID-19 patients. Non-critical patients n = 26; critical patients n = 36.