Vitamin D modulates systemic inflammation in patients with severe COVID-19

Aims The ability of vitamin D (VitD) to modulate immune responses in the clinical setting of COVID-19 infection is not well investigated. This study aimed to evaluate the ability of VitD to attenuate inflammatory responses in patients with severe COVID-19. Materials and methods Blood samples and nasopharyngeal swabs were obtained from patients with severe COVID-19 who had been treated (20 patients), or not (25 patients), with VitD, during their stay in the intensive care unit. Western blotting was used to evaluate the expressions of STAT3, JNK and AKT signaling pathways and ELISA was used to measure levels of IL-6, IL-17, and IL-1β in blood of these patients. Key findings Reduced levels of STAT3, JNK and AKT pathways and lower levels of proinflammatory cytokines such as IL-6, IL-17, and IL-1β were observed in VitD treated patients (50,000 IU of cholecalciferol weekly for 3 weeks), and in vitro following treatment of poly I:C stimulated PBMCs with VitD (50 nM of calcitriol). Moreover, lower circulatory levels of these proinflammatory cytokines following treatment with VitD were associated with lower serum levels of COVID-19-related severity markers such as D-dimer and C-reactive proteins (

Increased blood immune regulatory cells in severe COVID-19 with autoantibodies to type I interferons

Abstract The hallmark of severe COVID-19 is an uncontrolled inflammatory response, resulting from poorly understood immunological dysfunction. While regulatory T (Treg) and B (Breg) cells, as the main elements of immune homeostasis, contribute to the control of hyperinflammation during COVID-19 infection, we hypothesized change in their levels in relation to disease severity and the presence of autoantibodies (auto-Abs) to type I IFNs. Cytometric analysis of blood of 62 COVID-19 patients with different severities revealed an increased proportion of conventional (cTreg; CD25+FoxP3+) and unconventional (uTreg; CD25-FoxP3+) Tregs, as well as the LAG3+ immune suppressive form of cTreg/uTreg, in the blood of severe COVID-19 cases compared to the milder, non-hospitalized cases. The increase in blood levels of cTreg/uTreg, but not LAG3+ cTreg/uTreg subtypes, was even higher among patients with severe COVID-19 and auto-Abs to type I IFNs. Regarding Bregs, compared to the milder, non-hospitalized cases, the proportion of IL-35+ and IL-10+ Bregs was elevated in the blood of severe COVID-19 patients, and to a higher extent in those with auto-Abs to type I IFNs. Moreover, blood levels of cTreg, LAG3+ cTreg/uTreg, and IL-35+ and IL-10+ Breg subtypes were associated with lower blood levels of proinflammatory cytokines such as IL-6, IL-17, TNFα, and IL-1β. Interestingly, patients who were treated with either tocilizumab and/or a high dose of Vitamin D had higher blood levels of these regulatory cells and better control of the proinflammatory cytokines. These observations suggest that perturbations in the levels of immunomodulatory Tregs and Bregs occur in COVID-19, especially in the presence of auto-Abs to type I IFNs

Study raw data.

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Higher IL-17 level in saliva of severe COVID-19 patients.

(A) IL-17 mRNA levels in whole blood of COVID-19 patients with different severities. (B) IL-17 protein levels in plasma of COVID-19 patients with different severities. (C and D) IL-17 protein levels in saliva of COVID-19 patients with different severities and the associated ROC (receiver operating characteristic curve). (E-G) Correlation of IL-17 saliva level with serum levels of D-dimer, CRP (C-reactive protein), and ferritin of these patients. (H-K) TNFα and IL-1β protein levels in saliva of COVID-19 patients with different severities, and the associated ROCs. Specimens were collected from the following patients with COVID-19 (asymptomatic (n = 67), mild/moderate (n = 81), and severe (n = 53), as well as healthy controls (n = 50). Statistical test: Regression models were adjusted for demographics (age, gender, body mass index), comorbidity (diabetes mellitus) and severity markers of COVID-19 (CRP, D-dimer, and ferritin). ns: Non-significant, * P<0.05, *** P<0.001.</p

Clinical parameters of COVID-19 patients in according to disease severity.

Clinical parameters of COVID-19 patients in according to disease severity.</p

Correlation between IL-17 expression level and Th-17 signaling related cytokines/chemokines such as TNFα, IL-1β, IFNγ, IL-6, IL-8, and CCL2 in lung autopsies of COVID-19 patients (n = 17 SARS-CoV-2 infected lung tissues; GSE150316).

Data show that IL-17 level in COVID-19’ lung autopsies positively correlate with levels of TNFα, IL-1β, IFNγ, IL-6, IL-8 and CCL2, but not CCL2. Statistical test: Pearson’s coefficient test with two-tailed p-value (PDF)</p

Increased IL-17 level in saliva of severe COVID-19 patients associated with higher need for mechanical ventilation and/or death by days 29.

Kaplan–Meier survival curves of the need for mechanical ventilation (A-C) and/or death (D-F), based on the IL-17, TNFα, and IL-1β cytokine levels in saliva of patients with severe COVID-19 (n = 53). Statistical test: Cox proportional models adjusted for patient’s demographics factors (age, gender, and body mass index), comorbidities (diabetes mellitus), and COVID-19 related severity serum markers (D-dimer, CRP, and ferritin), with significance indicated by P value of less than 0.05.</p

Correlation between IL-17 expression level and Th-17 signaling related cytokines/chemokines such as TNFα, IL-1β, IFNγ, IL-6, neutrophils chemoattractant IL-8, and monocytes chemoattractant, CCL2 in nasopharyngeal swabs of COVID-19 patients (n = 430 COVID-19 patients; GSE152075).

Data show that IL-17 in these COVID-19’s nasopharyngeal swabs positively correlate with levels of IL-1β, TNFα, IL-6, IL-8 and CCL2, but not IFNγ. Statistical test: Pearson’s coefficient test with two-tailed p-value (PDF)</p

Increased IL-17 gene expression levels in lung and nasopharyngeal swabs of COVID-19 patients.

(A) IL-17 mRNA levels in nasopharyngeal swabs of COVID-19 patients compared to healthy controls (n = 430 COVID-19 patients vs n = 54 healthy controls; GSE152075). (B) IL-17 mRNA levels in lung autopsies of COVID-19 patients compared to healthy controls (n = 17 SARS-CoV-2 infected lung vs. n = 5 healthy lung biopsies; GSE150316). Statistical test: Unpaired t-test or Mann-Whitney U test, depending on the skewness of the data, * P (PDF)</p