Multi-omic and comparative analyses revealed monocyte-derived alpha-defensin-1 correlated with COVID-19 severity and inhibited SARS-CoV-2 infection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the etiological pathogen of coronavirus disease 2019 (COVID-19), a highly contagious disease, spreading quickly and threatening global public health. The symptoms of COVID-19 vary from mild reactions to severe respiratory distress or even fatal outcomes probably due to the different status of host immunity against the virus. Here in the study, we unveiled plasma proteomic signatures and transcriptional patterns of peripheral blood mononuclear cells (PBMCs) using blood samples of 10 COVID-19 patients with different severity. Through systemic analysis, α-defensin-1 (DEFA1) was identified to be elevated in both plasma and PBMCs, and correlated with disease severity and stages. In vitro study demonstrated that DEFA1 was secreted from immunocytes and suppressed SARS-CoV-2 infection of both original and mutated strains with dose dependency. By using sequencing data, we discovered that DEFA1 was activated in monocytes through NF-κB signaling pathway after infection, and secreted into circulation to perturb SARS-CoV-2 infection by interfering protein kinase C expression. It worked mainly during virus replication instead of entry in host cells. Together, the anti-SARS-CoV-2 mechanism of DEFA1 has unveiled a corner of how innate immunity is against SARS-CoV-2 and explored its clinical potential in disease prognosis and therapeutic intervention.

Diagnostic and Therapeutic Potential for HNP-1, HBD-1 and HBD-4 in Pregnant Women with COVID-19.

Pregnancy is characterized by significant immunological changes and a cytokine profile, as well as vitamin deficiencies that can cause problems for the correct development of a fetus. Defensins are small antimicrobial peptides that are part of the innate immune system and are involved in several biological activities. Following that, this study aims to compare the levels of various cytokines and to investigate the role of defensins between pregnant women with confirmed COVID-19 infection and pregnant women without any defined risk factor. TNF-α, TGF-ß, IL-2 and IL-10, ß-defensins, have been evaluated by gene expression in our population. At the same time, by ELISA assay IL-6, IL-8, defensin alpha 1, defensin beta 1 and defensin beta 4 have been measured. The data obtained show that mothers affected by COVID-19 have an increase in pro-inflammatory factors (TNF-α, TGF-ß, IL-2, IL-6, IL-8) compared to controls; this increase could generate a sort of "protection of the fetus" from virus attacks. Contemporarily, we have an increase in the anti-inflammatory cytokine IL-10 and an increase in AMPs, which highlights how the mother's body is responding to the viral attack. These results allow us to hypothesize a mechanism of "trafficking" of antimicrobial peptides from the mother to the fetus that would help the fetus to protect itself from the infection in progress.

Elevated Levels of Neutrophil Activated Proteins, Alpha-Defensins (DEFA1), Calprotectin (S100A8/A9) and Myeloperoxidase (MPO) Are Associated With Disease Severity in COVID-19 Patients.

Understanding of the basis for severity and fatal outcome of SARS-CoV-2 infection is of paramount importance for developing therapeutic options and identification of prognostic markers. So far, accumulation of neutrophils and increased levels of pro-inflammatory cytokines are associated with disease severity in COVID-19 patients. In this study, we aimed to compare circulatory levels of neutrophil secretory proteins, alpha-defensins (DEFA1), calprotectin (S100A8/A9), and myeloperoxidase (MPO) in COVID-19 patients with different clinical presentations. We studied 19 healthy subjects, 63 COVID-19 patients with mild (n=32) and severe (n=31) disease, 23 asymptomatic individuals identified through contact tracing programme and 23 recovering patients (1-4 months post-disease). At the time of disease presentation, serum levels of DEFA1 were significantly higher in patients with mild (mean230 ± 17, p<0.0001) and severe (mean452 ± 46, p<0.0001) disease respectively in comparison to healthy subjects (mean113 ± 11). S100A8/A9 proteins were significantly higher in COVID-19 patients (p<0.0001) irrespective of disease severity. The levels of DEFA1, S100A8/A9 and MPO reduced to normal in recovering patients and comparable to healthy subjects. Surprisingly, DEFA1 levels were higher in severe than mild patients in first week of onset of disease (p=0.004). Odds-ratio analysis showed that DEFA1 could act as potential biomarker in predicting disease severity (OR=11.34). In addition, levels of DEFA1 and S100A8/A9 were significantly higher in patients with fatal outcome (p=0.004 and p=0.03) respectively. The rise in DEFA1 levels was independent of secondary infections. In conclusion, our data suggest that induction of elevated levels of alpha-defensins and S100A8/A9 is associated with poor disease outcome in COVID-19 patients.

Divergent impacts of tocilizumab and colchicine in COVID-19-associated coagulopathy: the role of alpha-defensins.

Patients who are severely affected by coronavirus disease 2019 (COVID-19) may develop a delayed onset 'cytokine storm', which includes an increase in interleukin-6 (IL-6). This may be followed by a pro-thrombotic state and increased D-dimers. It was anticipated that tocilizumab (TCZ), an anti-IL-6 receptor monoclonal antibody, would mitigate inflammation and coagulation in patients with COVID-19. However, clinical trials with TCZ have recorded an increase in D-dimer levels. In contrast to TCZ, colchicine reduced D-dimer levels in patients with COVID-19. To understand how the two anti-inflammatory agents have diverse effects on D-dimer levels, we present data from two clinical trials that we performed. In the first trial, TCZ was administered (8 mg/kg) to patients who had a positive polymerase chain reaction test for COVID-19. In the second trial, colchicine was given (0·5 mg twice a day). We found that TCZ significantly increased IL-6, α-Defensin (α-Def), a pro-thrombotic peptide, and D-dimers. In contrast, treatment with colchicine reduced α-Def and Di-dimer levels. In vitro studies show that IL-6 stimulated the release of α-Def from human neutrophils but in contrast to colchicine, TCZ did not inhibit the stimulatory effect of IL-6; raising the possibility that the increase in IL-6 in patients with COVID-19 treated with TCZ triggers the release of α-Def, which promotes pro-thrombotic events reflected in an increase in D-dimer levels.

Inhibition of SARS-CoV-2 Infection by Human Defensin HNP1 and Retrocyclin RC-101.

Severe acute respiratory syndrome coronavirus (SARS-CoV)-2 is an enveloped virus responsible for the COVID-19 pandemic. The emergence of new potentially more transmissible and vaccine-resistant variants of SARS-CoV-2 is an ever-present threat. Thus, it remains essential to better understand innate immune mechanisms that can inhibit the virus. One component of the innate immune system with broad antipathogen, including antiviral, activity is a group of cationic immune peptides termed defensins. The ability of defensins to neutralize enveloped and non-enveloped viruses and to inactivate numerous bacterial toxins correlate with their ability to promote the unfolding of proteins with high conformational plasticity. We found that human neutrophil α-defensin HNP1 binds to SARS-CoV-2 Spike protein with submicromolar affinity that is more than 20 fold stronger than its binding to serum albumin. As such, HNP1, as well as a θ-defensin retrocyclin RC-101, both interfere with Spike-mediated membrane fusion, Spike-pseudotyped lentivirus infection, and authentic SARS-CoV-2 infection in cell culture. These effects correlate with the abilities of the defensins to destabilize and precipitate Spike protein and inhibit the interaction of Spike with the ACE2 receptor. Serum reduces the anti-SARS-CoV-2 activity of HNP1, though at high concentrations, HNP1 was able to inactivate the virus even in the presence of serum. Overall, our results suggest that defensins can negatively affect the native conformation of SARS-CoV-2 Spike, and that α- and θ-defensins may be valuable tools in developing SARS-CoV-2 infection prevention strategies.

Alpha-defensins: risk factor for thrombosis in COVID-19 infection.

The inflammatory response to SARS/CoV-2 (COVID-19) infection may contribute to the risk of thromboembolic complications. α-Defensins, antimicrobial peptides released from activated neutrophils, are anti-fibrinolytic and prothrombotic in vitro and in mouse models. In this prospective study of 176 patients with COVID-19 infection, we found that plasma levels of α-defensins were elevated, tracked with disease progression/mortality or resolution and with plasma levels of interleukin-6 (IL-6) and D-dimers. Immunohistochemistry revealed intense deposition of α-defensins in lung vasculature and thrombi. IL-6 stimulated the release of α-defensins from neutrophils, thereby accelerating coagulation and inhibiting fibrinolysis in human blood, imitating the coagulation pattern in COVID-19 patients. The procoagulant effect of IL-6 was inhibited by colchicine, which blocks neutrophil degranulation. These studies describe a link between inflammation and the risk of thromboembolism, and they identify a potential new approach to mitigate this risk in patients with COVID-19 and potentially in other inflammatory prothrombotic conditions.

Evaluation of alpha defensin, IL-1 receptor antagonist, and IL-18 levels in COVID-19 patients with macrophage activation syndrome and acute respiratory distress syndrome.

BACKGROUND: Many laboratory parameters have been associated with morbidity and mortality in SARS-CoV-2 (COVID-19), which emerged in an animal market in Wuhan, China in December 2019 and has infected over 20 million people. This study investigated the relationship between serum interleukin (IL)-18, IL-1 receptor antagonist (IL-1Ra), and alpha defensin levels and the clinical course and prognosis of COVID-19. MATERIALS AND METHODS: This study included 100 patients who were admitted to the chest diseases department and intensive care unit of our hospital and diagnosed with COVID-19 by real-time polymerase chain reaction (PCR) of nasopharyngeal swab samples between March 24 and May 31, 2020. The control group consisted of 50 nonsymptomatic health workers with negative real-time PCR results in routine COVID-19 screening in our hospital. RESULTS: Serum alpha defensin, IL-1Ra, and IL-18 levels were significantly higher in patients who developed macrophage activation syndrome (MAS) and acute respiratory distress syndrome (ARDS) compared to patients who did not (p < .001 for all). Alpha defensin, IL-1Ra, and IL-18 levels were significantly higher in COVID-19 patients with and without MAS or ARDS when compared to the control group (p < .001 for all). When the 9 patients who died were compared with the 91 surviving patients, IL-1Ra and IL-18 levels were found to be significantly higher in the nonsurvivors (p < .001). CONCLUSION: Our findings of correlations between alpha defensin and levels of IL-1Ra and IL-18, which were previously shown to be useful in COVID-19 treatment and follow-up, indicates that it may also be promising in treatment.

Defensin 5 for prevention of SARS-CoV-2 invasion and Covid-19 disease.

Corona virus disease 2019 (Covid-19), a pandemia emerged recently, caused by severe acute respiratory syndrome corona virus 2 (SARS-CoV-2). The receptor for corona virus and influenza A is the mucosal cell membrane protein angiotensin converting enzyme 2 (ACE2), which is abundant on the membrane of alveolar cells and enterocytes. Viral spike protein 1 (S1) is the ligand, with an affinity of 14.7 nM to the receptor. The main port of entry for the virus is the upper respiratory tract, and the diagnosis is usually by PCR of the viral RNA with nasal and pharyngeal swab test. Human defensin 5 (HDEF5) is a protein encoded by the DEFA gene, secreted by Paneth cells in the small intestine and by granules of neutrophils. It has an affinity of 39.3 nM to ACE2, much higher than that of the corona S1. HDEF5 may also attach to glycosylated Corona S1 protein, make its efficiency even better. The issues to be investigated are the affinity of HDEF5 to S1 protein, the ability of recombinant HDEF5 function in attaching both ACE2 and S1, and the feasibility to perform aerosol spray of this protein. In addition, safety and efficiency should be studied in phases I, II and II clinical protocols. Thus, an aerosol spray of HDEF5 given through the nose and throat, once to several times a day, may be a very efficient approach to prevent infection with SARA-CoV-2 as well as influenza A.