ABSTRACT
Background: Evidence suggest that critically ill COVID-19 patients have an impairment of IFN-I response. This defect in antiviral mechanisms is explained in some patients by the presence of anti IFN-alpha neutralizing autoantibodies (NAbs). However, whether NAbs fluctuate longitudinally during COVID-19, and what are their specificity toward IFN-I subtypes and consequences on the IFN response remain elusive. Methods: Binding antibodies (BAbs) to IFN-alpha and IFN-beta were screened in serum samples (n=360) of COVID-19 patients using ELISA assays. All serum samples containing BAbs were processed to investigate NAbs using antiviral bioassay. Respiratory samples (n=17) were also included for the NAbs analysis. Transcript levels of IFN-alpha, IFN-beta, IFN-omega and IFN stimulated genes (ISGs) were analyzed through RT/Real Time PCR. Results: Results showed that 16.94% (61/360) of COVID-19 patients had circulating BAbs against IFN-alpha and IFN-beta. Further, 21% (13/61) of critically ill subjects had NAbs with a variable titer against all the IFN-alpha subtypes (70-71680 TRU/ml) while only 1 patient had anti IFN-beta NAbs. About 70% of these serum samples showed cross reactivity to IFN-omega at different extent (27-106667 TRU/ml). Longitudinal evaluation at different time points after hospitalization indicate the persistence of high NAbs titer throughout the time. NAbs to IFN-alpha (10-20 TRU/ml) were also detected in 17.64% of respiratory samples. Patients with NAbs had severe disease and exhibited alterations in the levels of many hematological indicators [(white blood cells, neutrophils, platelets, neutrophils to lymphocytes ratio, platelets to lymphocytes ratio, D-dimer, C-reactive protein and lactate dehydrogenase;p<0.05)]. Transcriptomic analysis indicated that levels of IFN genes were lower in NAbs patients than in healthy donors (p≤0.05). However, only the ISGs levels were reduced compared to those found in the NAbs negative patients. Of note, expression of ISGs, was abolished during hospitalization in all patients with persistent high titer of NAbs. Conclusion: Our finding demonstrate that NAbs with a broad specificity to IFN-I can be found in blood and respiratory samples from severe COVID-19 patients. NAbs detection was associated with a defective IFN response and with an increased levels of markers of disease severity.
ABSTRACT
Oral bacteriotherapy with SAB51 has been previously evidenced as support of standard care in treating patients hospitalized for SARS-CoV2 pneumonia and associated gastrointestinal symptoms leading to a significantly decreased risk of developing respiratory failure. This study aimed to compare the rate of mortality, the need for ICU hospitalization, and the length of hospitalization in patients with severe COVID-19 pneumonia who received the best available therapy in Italy (BAT) versus patients treated with BAT and supplemented with SLAB51. 200 adults (≥18 years) hospitalized from March 6 until April 26, 2020 with severe COVID-19 pneumonia and needing oxygen support were investigated. All patients received therapeutic regimens including hydroxychloroquine (200 mg twice a day for 7 days), azithromycin (500 mg once a day for 7 days), lopinavir–ritonavir (400/100 mg twice a day) or darunavir–cobicistat (800/150 mg once a day) for 14 days, and low molecular weight heparin. Out of the 200 patients, 112 received BAT without oral bacteriotherapy and 88 BAT with SLAB51. Crude mortality was 22%. Significantly higher mortality has been observed in the group of patients managed only with BAT (30%) to that treated with BAT plus SLAB51 (11%). In both groups, the highest mortality was registered for the combination of Tocilizumab and hydroxychloroquine and Tocilizumab. By multivariate analysis, the age >65 years, PCR >41.8 mg/L, Platelets <150.000 mmc, and CV events were associated with increased mortality risk. Overall results evidenced that oral bacteriotherapy with SLAB51 was an independent variable associated with a reduced risk for death.(Image presented) Death probability. Kaplan-Meier curves displaying Best Available therapy (BAT) versus BAT and oral bacteriotherapy.
ABSTRACT
Background: A severe SARS-CoV-2 related immunopathology may be the driver cause underlying the deleterious clinical manifestations observed in COVID-19 patients. To identify possible tissue-specific immune responses patterns, a compartmental immunophenotyping analysis of CD4+ and CD8+ T lymphocytes and IFN response has been performed in SARS-CoV-2 infected subjects with acute respiratory distress syndrome. Methods: Bronchoalveolar lavage (BAL) and Peripheral Blood Mononuclear Cells (PBMC) samples were collected from 13 SARS-CoV-2 infected subjects (9 males and 4 females) consecutively admitted to intensive care unit (ICU) of Policlinico Umberto I, Sapienza University Hospital in Rome (Italy). The frequencies of CD4+, CD8+ T lymphocytes and those expressing immune activation markers (CD38, HLADR), naïve, central memory (CMEM), and effector memory (TEM) T cell subsets were evaluated in both anatomical sites by multiparametric flow cytometry. Gene expression levels of Interferon regulatory factor 7 (IRF7) and the Interferon Stimulated Gene 15 (ISG15) were evaluated in BAL and PBMC by Real-time PCR. Results: Critically SARS-CoV-2 infected patients exhibited a lung compartmentalization of CD8+ T cells (p=0.003), with a lower CD4/CD8 ratio in BAL compared to blood district (p<0.01). However, higher frequencies of CD8+ T cells were recorded in PBMC of female SARS-CoV-2 infected patients (p=0.04) and the same trend was observed in the lung compartment. By contrast, a trend of increasing CD4+ T cells frequencies was observed in BAL samples of male patients, as opposed to blood compartment. Additionally, an increased expression of immune activation markers CD38 and HLADR has been detected in BAL CD8+ T cells (p<0.01) as well as in blood CD4+ T cells (p=0.03). An increased frequency of CD4+ and CD8+ TEM cells has been documented in BAL of SARS-CoV-2 infected patients (p<0.05), as opposed to higher frequencies of CD4+ and CD8+ TCM cells recorded in the blood compartment (p<0.01). Notably, higher levels of ISG15 and IRF7 found in BAL were inversely associated to activated CD8+ T cell frequencies in the lung compartment compared to blood district (ISG15: r=-0.570, p<0.05) (IRF7: r=-0.683, p=0.01). Conclusion: Our findings provide new insight into a distinct T cells profile and IFN genes expression in the lung and in the blood compartment of SARS-CoV-2 infected patients, that might be highly relevant for the clinical course of COVID-19.