COVID-19 adenovirus vector vaccine induces higher interferon and pro-inflammatory responses than mRNA vaccines in human PBMCs, macrophages and moDCs

Background During the COVID-19 pandemic multiple vaccines were rapidly developed and widely used throughout the world. At present there is very little information on COVID-19 vaccine interactions with primary human immune cells such as peripheral blood mononuclear cells (PBMCs), monocyte-derived macrophages and dendritic cells (moDCs). Methods Human PBMCs, macrophages and moDCs were stimulated with different COVID-19 vaccines, and the expression of interferon (IFN-λ1, IFN-α1), pro-inflammatory (IL-1β, IL-6, IL-8, IL-18, CXCL-4, CXCL-10, TNF-α) and Th1-type cytokine mRNAs (IL-2, IFN-γ) were analyzed by qPCR. In addition, the expression of vaccine induced spike (S) protein and antiviral molecules were studied in primary immune cells and in A549 lung epithelial cells. Results Adenovirus vector (Ad-vector) vaccine AZD1222 induced high levels of IFN-λ1, IFN-α1, CXCL-10, IL-6, and TNF-α mRNAs in PBMCs at early time points of stimulation while the expression of IFN-γ and IL-2 mRNA took place at later times. AZD1222 also induced IFN-λ1, CXCL-10 and IL-6 mRNA expression in monocyte-derived macrophages and DCs in a dose-dependent fashion. AZD1222 also activated the phosphorylation of IRF3 and induced MxA expression. BNT162b2 and mRNA-1273 mRNA vaccines failed to induce or induced very weak cytokine gene expression in all cell models. None of the vaccines enhanced the expression of CXCL-4. AZD1222 and mRNA-1273 vaccines induced high expression of S protein in all studied cells. Conclusions Ad-vector vaccine induces higher IFN and pro-inflammatory responses than the mRNA vaccines in human immune cells. This data shows that AZD1222 readily activates IFN and pro-inflammatory cytokine gene expression in PBMCs, macrophages and DCs, but fails to further enhance CXCL-4 mRNA expression.

Th-1, Th-2, Th-9, Th-17, Th-22 type cytokine concentrations of critical COVID-19 patients after treatment with Remdesivir.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) rapidly spread around the world causing a pandemic known as coronavirus disease 2019 (COVID-19). Cytokine storm was directly correlated with severity of COVID-19 syndromes. We evaluated the levels of 13 cytokines in ICU hospitalized COVID-19 patients (n = 29) before, and after treatment with Remdesivir as well as in healthy controls (n = 29). Blood samples were obtained from ICU patients during ICU admission (before treatment) and 5 days after treatment with Remdesivir. A group of 29 age- and gender-matched healthy controls was also studied. Cytokine levels were evaluated by multiplex immunoassay method using a fluorescence labeled cytokine panel. In comparison to cytokine levels measured at ICU admission, serum levels were reduced of IL-6 (134.75 pg/mL vs. 20.73 pg/mL, P < 0.0001), TNF-α (121.67 pg/mL vs. 10.15 pg/mL, P < 0.0001) and IFN-γ (29.69 pg/mL vs. 22.27 pg/mL, P = 0.005), whereas serum level was increased of IL-4 (8.47 pg/mL vs. 12.44 pg/mL, P = 0.002) within 5 days after Remdesivir treatment. Comparing with before treatment, Remdesivir significantly reduced the levels of inflammatory (258.98 pg/mL vs. 37.43 pg/mL, P < 0.0001), Th1-type (31.24 pg/mL vs. 24.46 pg/mL, P = 0.007), and Th17-type (36.79 pg/mL vs. 26.22 pg/mL, P < 0.0001) cytokines in critical COVID-19 patients. However, after Remdesivir treatment, the concentrations of Th2-type cytokines were significantly higher than before treatment (52.69 pg/mL vs. 37.09 pg/mL, P < 0.0001). In conclusion, Remdesivir led to decrease levels of Th1-type and Th17-type cytokines and increase Th2-type cytokines in critical COVID-19 patients 5 days after treatment.

Intranasal administration of adenoviral vaccines expressing SARS-CoV-2 spike protein improves vaccine immunity in mouse models.

The ongoing SARS-CoV-2 pandemic is controlled but not halted by public health measures and mass vaccination strategies which have exclusively relied on intramuscular vaccines. Intranasal vaccines can prime or recruit to the respiratory epithelium mucosal immune cells capable of preventing infection. Here we report a comprehensive series of studies on this concept using various mouse models, including HLA class II-humanized transgenic strains. We found that a single intranasal (i.n.) dose of serotype-5 adenoviral vectors expressing either the receptor binding domain (Ad5-RBD) or the complete ectodomain (Ad5-S) of the SARS-CoV-2 spike protein was effective in inducing i) serum and bronchoalveolar lavage (BAL) anti-spike IgA and IgG, ii) robust SARS-CoV-2-neutralizing activity in the serum and BAL, iii) rigorous spike-directed T helper 1 cell/cytotoxic T cell immunity, and iv) protection of mice from a challenge with the SARS-CoV-2 beta variant. Intramuscular (i.m.) Ad5-RBD or Ad5-S administration did not induce serum or BAL IgA, and resulted in lower neutralizing titers in the serum. Moreover, prior immunity induced by an intramuscular mRNA vaccine could be potently enhanced and modulated towards a mucosal IgA response by an i.n. Ad5-S booster. Notably, Ad5 DNA was found in the liver or spleen after i.m. but not i.n. administration, indicating a lack of systemic spread of the vaccine vector, which has been associated with a risk of thrombotic thrombocytopenia. Unlike in otherwise genetically identical HLA-DQ6 mice, in HLA-DQ8 mice Ad5-RBD vaccine was inferior to Ad5-S, suggesting that the RBD fragment does not contain a sufficient collection of helper-T cell epitopes to constitute an optimal vaccine antigen. Our data add to previous promising preclinical results on intranasal SARS-CoV-2 vaccination and support the potential of this approach to elicit mucosal immunity for preventing transmission of SARS-CoV-2.

Th-1, Th-2, Th-9, Th-17, Th-22 type cytokine concentrations of critical COVID-19 patients after treatment with Remdesivir

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) rapidly spread around the world causing a pandemic known as coronavirus disease 2019 (COVID-19). Cytokine storm was directly correlated with severity of COVID-19 syndromes. We evaluated the levels of 13 cytokines in ICU hospitalized COVID-19 patients (n=29) before, and after treatment with Remdesivir as well as in healthy controls (n=29). Blood samples were obtained from ICU patients during ICU admission (before treatment) and 5 days after treatment with Remdesivir. A group of 29 age- and gender-matched healthy controls was also studied. Cytokine levels were evaluated by multiplex immunoassay method using a fluorescence labeled cytokine panel. In comparison to cytokine levels measured at ICU admission, serum levels were reduced of IL-6 (134.75 pg/mL vs. 20.73 pg/mL, P< 0.0001), TNF-α (121.67 pg/mL vs. 10.15 pg/mL, P< 0.0001) and IFN-γ (29.69 pg/mL vs. 22.27 pg/mL, P= 0.005), whereas serum level was increased of IL-4 (8.47 pg/mL vs. 12.44 pg/mL, P= 0.002) within 5 days after Remdesivir treatment. Comparing with before treatment, Remdesivir significantly reduced the levels of inflammatory (258.98 pg/mL vs. 37.43 pg/mL, P< 0.0001), Th1-type (31.24 pg/mL vs. 24.46 pg/mL, P= 0.007), and Th17-type (36.79 pg/mL vs. 26.22 pg/mL, P< 0.0001) cytokines in critical COVID-19 patients. However, after Remdesivir treatment, the concentrations of Th2-type cytokines were significantly higher than before treatment (52.69 pg/mL vs. 37.09 pg/mL, P< 0.0001). In conclusion, Remdesivir led to decrease levels of Th1-type and Th17-type cytokines and increase Th2-type cytokines in critical COVID-19 patients 5 days after treatment.

Immune responses in mildly versus critically ill COVID-19 patients.

The current coronavirus pandemic (COVID-19), caused by SARS-CoV-2, has had devastating effects on the global health and economic system. The cellular and molecular mediators of both the innate and adaptive immune systems are critical in controlling SARS-CoV-2 infections. However, dysregulated inflammatory responses and imbalanced adaptive immunity may contribute to tissue destruction and pathogenesis of the disease. Important mechanisms in severe forms of COVID-19 include overproduction of inflammatory cytokines, impairment of type I IFN response, overactivation of neutrophils and macrophages, decreased frequencies of DC cells, NK cells and ILCs, complement activation, lymphopenia, Th1 and Treg hypoactivation, Th2 and Th17 hyperactivation, as well as decreased clonal diversity and dysregulated B lymphocyte function. Given the relationship between disease severity and an imbalanced immune system, scientists have been led to manipulate the immune system as a therapeutic approach. For example, anti-cytokine, cell, and IVIG therapies have received attention in the treatment of severe COVID-19. In this review, the role of immunity in the development and progression of COVID-19 is discussed, focusing on molecular and cellular aspects of the immune system in mild vs. severe forms of the disease. Moreover, some immune- based therapeutic approaches to COVID-19 are being investigated. Understanding key processes involved in the disease progression is critical in developing therapeutic agents and optimizing related strategies.

Immune responses in mildly versus critically ill COVID-19 patients

The current coronavirus pandemic (COVID-19), caused by SARS-CoV-2, has had devastating effects on the global health and economic system. The cellular and molecular mediators of both the innate and adaptive immune systems are critical in controlling SARS-CoV-2 infections. However, dysregulated inflammatory responses and imbalanced adaptive immunity may contribute to tissue destruction and pathogenesis of the disease. Important mechanisms in severe forms of COVID-19 include overproduction of inflammatory cytokines, impairment of type I IFN response, overactivation of neutrophils and macrophages, decreased frequencies of DC cells, NK cells and ILCs, complement activation, lymphopenia, Th1 and Treg hypoactivation, Th2 and Th17 hyperactivation, as well as decreased clonal diversity and dysregulated B lymphocyte function. Given the relationship between disease severity and an imbalanced immune system, scientists have been led to manipulate the immune system as a therapeutic approach. For example, anti-cytokine, cell, and IVIG therapies have received attention in the treatment of severe COVID-19. In this review, the role of immunity in the development and progression of COVID-19 is discussed, focusing on molecular and cellular aspects of the immune system in mild vs. severe forms of the disease. Moreover, some immune- based therapeutic approaches to COVID-19 are being investigated. Understanding key processes involved in the disease progression is critical in developing therapeutic agents and optimizing related strategies.

Impaired immunity and high attack rates caused by SARS-CoV-2 variants among vaccinated long-term care facility residents.

INTRODUCTION: Long-term care facilities (LTCF) residents are at high risk for severe coronavirus disease 2019 (COVID-19), and therefore, COVID-19 vaccinations were prioritized for residents and personnel in Finland at the beginning of 2021. METHODS: We investigated COVID-19 outbreaks in two LTCFs, where residents were once or twice vaccinated. After the outbreaks we measured immunoglobulin G (IgG) antibodies to severe acute respiratory syndrome coronavirus 2 spike glycoprotein, neutralizing antibody (NAb) titers, and cell-mediated immunity markers from residents and healthcare workers (HCWs). RESULTS: In LTFC-1, the outbreak was caused by an Alpha variant (B.1.1.7) and the attack rate (AR) among once vaccinated residents was 23%. In LTCF-2 the outbreak was caused by a Beta variant (B.1.351). Its AR was 47% although all residents had received their second dose 1 month before the outbreak. We observed that vaccination had induced lower IgG concentrations, NAb titers and cell-mediated immune responses in residents compared to HCWs. Only 1/8 residents had NAb to the Beta variant after two vaccine doses. CONCLUSIONS: The vaccinated elderly remain susceptible to breakthrough infections caused by Alpha and Beta variants. The weaker vaccine response in the elderly needs to be addressed in vaccination protocols, while new variants capable of evading vaccine-induced immunity continue to emerge.

Impaired immunity and high attack rates caused by SARS‐CoV‐2 variants among vaccinated long‐term care facility residents

Introduction Long‐term care facilities (LTCF) residents are at high risk for severe coronavirus disease 2019 (COVID‐19), and therefore, COVID‐19 vaccinations were prioritized for residents and personnel in Finland at the beginning of 2021. Methods We investigated COVID‐19 outbreaks in two LTCFs, where residents were once or twice vaccinated. After the outbreaks we measured immunoglobulin G (IgG) antibodies to severe acute respiratory syndrome coronavirus 2 spike glycoprotein, neutralizing antibody (NAb) titers, and cell‐mediated immunity markers from residents and healthcare workers (HCWs). Results In LTFC‐1, the outbreak was caused by an Alpha variant (B.1.1.7) and the attack rate (AR) among once vaccinated residents was 23%. In LTCF‐2 the outbreak was caused by a Beta variant (B.1.351). Its AR was 47% although all residents had received their second dose 1 month before the outbreak. We observed that vaccination had induced lower IgG concentrations, NAb titers and cell‐mediated immune responses in residents compared to HCWs. Only 1/8 residents had NAb to the Beta variant after two vaccine doses. Conclusions The vaccinated elderly remain susceptible to breakthrough infections caused by Alpha and Beta variants. The weaker vaccine response in the elderly needs to be addressed in vaccination protocols, while new variants capable of evading vaccine‐induced immunity continue to emerge. IgG levels to WT SARS‐CoV‐2 spike glycoprotein after vaccination with Comirnaty. (A) Receptor binding domain (RBD) and (B) full‐length spike glycoprotein (SFL) in binding antibody units (BAU)/ml. Dashed lines mark the threshold for positive result per antigen. Statistical significance measured with Wilcoxon rank‐sum test, significance level 0.05.

Common Laboratory Mice Are Susceptible to Infection with the SARS-CoV-2 Beta Variant.

Small animal models are of crucial importance for assessing COVID-19 countermeasures. Common laboratory mice would be well-suited for this purpose but are not susceptible to infection with wild-type SARS-CoV-2. However, the development of mouse-adapted virus strains has revealed key mutations in the SARS-CoV-2 spike protein that increase infectivity, and interestingly, many of these mutations are also present in naturally occurring SARS-CoV-2 variants of concern. This suggests that these variants might have the ability to infect common laboratory mice. Herein we show that the SARS-CoV-2 beta variant attains infectibility to BALB/c mice and causes pulmonary changes within 2-3 days post infection, consistent with results seen in other murine models of COVID-19, at a reasonable virus dose (2 × 105 PFU). The findings suggest that common laboratory mice can serve as the animal model of choice for testing the effectiveness of antiviral drugs and vaccines against SARS-CoV-2.

COVID-19 adenovirus vaccine triggers antibodies against PF4 complexes to activate complement and platelets.

BACKGROUND: Vaccine-induced thrombotic thrombocytopenia (VITT) is a rare coagulation disorder reported after administration of COVID-19 adenovirus-vectored vaccines. VITT is mediated by anti-platelet factor 4 (PF4) antibodies activating platelets through the Fcγ-receptor II (FcγRII), and it is associated with strong fibrin turnover. The complement system is involved in several other immunothrombotic entities, but its impact on VITT is not established. OBJECTIVE: To assess antibodies in interaction with the activation of platelets and complement triggered by VITT. METHODS: Antibodies against adenovirus type 2 hexon protein, ChAdOx1 adenoviral vector-specific IgG and PF4 were analyzed by enzyme immunoassays from VITT patients (n = 5). The EDTA plasma samples of the patients and controls were used to measure both terminal complement complexes (TCC) by ELISA and aggregation of healthy donor platelets. We studied the effects of human immunoglobulin (IVIG) and glycoprotein IIb/IIIa inhibitor (GPIIb/IIIa) on spontaneous and collagen-induced platelet aggregation supplemented with VITT plasma. RESULTS: None of the patients had experienced a COVID-19 infection. Antibody analyses confirmed the immunogenicity of the adenovirus-vectored ChAdOx1 vaccine. Moreover, VITT plasma had anti-PF4 antibodies and elevated TCC levels as a sign of complement activation. In isolated healthy donor platelets, VITT patient plasma caused marked, spontaneous aggregation of platelets, which was abolished by eptifibatide and high-dose therapeutic IVIG. CONCLUSIONS: Our findings suggest that VITT is triggered by antibodies against adenovirus vector and PF4-polyanion complexes which strongly co-activate complement and platelets. The spontaneous platelet aggregation was suppressed by IVIG or eptifibatide, indicating that besides FcγRII, also GPIIb/IIIa receptor exerts platelet procoagulant role in VITT.

The Effects of Type 2 Diabetes Mellitus on Organ Metabolism and the Immune System.

Metabolic abnormalities such as dyslipidemia, hyperinsulinemia, or insulin resistance and obesity play key roles in the induction and progression of type 2 diabetes mellitus (T2DM). The field of immunometabolism implies a bidirectional link between the immune system and metabolism, in which inflammation plays an essential role in the promotion of metabolic abnormalities (e.g., obesity and T2DM), and metabolic factors, in turn, regulate immune cell functions. Obesity as the main inducer of a systemic low-level inflammation is a main susceptibility factor for T2DM. Obesity-related immune cell infiltration, inflammation, and increased oxidative stress promote metabolic impairments in the insulin-sensitive tissues and finally, insulin resistance, organ failure, and premature aging occur. Hyperglycemia and the subsequent inflammation are the main causes of micro- and macroangiopathies in the circulatory system. They also promote the gut microbiota dysbiosis, increased intestinal permeability, and fatty liver disease. The impaired immune system together with metabolic imbalance also increases the susceptibility of patients to several pathogenic agents such as the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Thus, the need for a proper immunization protocol among such patients is granted. The focus of the current review is to explore metabolic and immunological abnormalities affecting several organs of T2DM patients and explain the mechanisms, whereby diabetic patients become more susceptible to infectious diseases.