Human Properdin Released By Infiltrating Neutrophils Can Modulate Influenza A Virus Infection

The complement system is designed to recognise and eliminate invading pathogens via activation of classical, alternative and lectin pathways. Human properdin stabilises the alternative pathway C3 convertase, resulting in an amplification loop that leads to the formation of C5 convertase, thereby acting as a positive regulator of the alternative pathway. It has been noted that human properdin on its own can operate as a pattern recognition receptor and exert immune functions outside its involvement in complement activation. Properdin can bind directly to microbial targets via DNA, sulfatides and glycosaminoglycans, apoptotic cells, nanoparticles, and well-known viral virulence factors. This study was aimed at investigating the complement-independent role of properdin against Influenza A virus infection. As one of the first immune cells to arrive at the site of IAV infection, we show here that IAV challenged neutrophils released properdin in a time-dependent manner. Properdin was found to directly interact with haemagglutinin, neuraminidase and matrix 1 protein Influenza A virus proteins in ELISA and western blot. Furthermore, modelling studies revealed that properdin could bind HA and NA of the H1N1 subtype with higher affinity compared to that of H3N2 due to the presence of an HA cleavage site in H1N1. In an infection assay using A549 cells, properdin suppressed viral replication in pH1N1 subtype while promoting replication of H3N2 subtype, as revealed by qPCR analysis of M1 transcripts. Properdin treatment triggered an anti-inflammatory response in H1N1-challenged A549 cells and a pro-inflammatory response in H3N2-infected cells, as evident from differential mRNA expression of TNF-α, NF-κB, IFN-α, IFN-β, IL-6, IL-12 and RANTES. Properdin treatment also reduced luciferase reporter activity in MDCK cells transduced with H1N1 pseudotyped lentiviral particles; however, it was increased in the case of pseudotyped H3N2 particles. Collectively, we conclude that infiltrating neutrophils at the site of IAV infection can release properdin, which then acts as an entry inhibitor for pandemic H1N1 subtype while suppressing viral replication and inducing an anti-inflammatory response. H3N2 subtype can escape this immune restriction due to altered haemagglutinin and neuraminindase, leading to enhanced viral entry, replication and pro-inflammatory response. Thus, depending on the subtype, properdin can either limit or aggravate IAV infection in the host.

Lung Neutrophilic Recruitment and IL-8/IL-17A Tissue Expression in COVID-19

The new SARS-CoV-2 virus differs from the pandemic Influenza A virus H1N1 subtype (H1N1pmd09) how it induces a pro-inflammatory response in infected patients. This study aims to evaluate the involvement of SNPs and tissue expression of IL-17A and the neutrophils recruitment in post-mortem lung samples from patients who died of severe forms of COVID-19 comparing to those who died by H1N1pdm09. Twenty lung samples from patients SARS-CoV-2 infected (COVID-19 group) and 10 lung samples from adults who died from a severe respiratory H1N1pdm09 infection (H1N1 group) were tested. The tissue expression of IL-8/IL-17A was identified by immunohistochemistry, and hematoxylin and eosin (H&E) stain slides were used for neutrophil scoring. DNA was extracted from paraffin blocks, and genotyping was done in real time-PCR for two IL17A target polymorphisms. Tissue expression increasing of IL-8/IL-17A and a higher number of neutrophils were identified in samples from the H1N1 group compared to the COVID-19 group. The distribution of genotype frequencies in the IL17A gene was not statistically significant between groups. However, the G allele (GG and GA) of rs3819025 was correlated with higher tissue expression of IL-17A in the COVID-19 group. SARS-CoV-2 virus evokes an exacerbated response of the host’s immune system but differs from that observed in the H1N1pdm09 infection since the IL-8/IL-17A tissue expression, and lung neutrophilic recruitment may be decreased. In SNP rs3819025 (G/A), the G allele may be considered a risk allele in the patients who died for COVID-19.

Alveolar Neutrophilic Recruitment in COVID-19 May Not be Mediated by Th17 Response

Introduction: The COVID-19 current pandemic disease differs from the H1N1pdm09 caused by Influenza A virus H1N1 subtype, by how it induces a pro-inflammatory response in infected lungs. Objective: Investigate the role of Th17 response in the pathogenesis of COVID-19 injury by analyzing the tissue expression of interleukins 8 and 17A and the neutrophils in lung samples of patients who die of COVID-19, comparing to H1N1pdm09. Study design and post-mortem results: Six lung samples from patients SARS-CoV-2 infected (COVID-19 group), and ten lung samples from adults who died from H1N1pdm09 (H1N1 group), were tested. A control group was also added to the study. H&E slides were used for neutrophils scoring. The tissue expression of IL-8 and IL-17A was identified by immunohistochemistry. Tissue expression increasing of IL-17A and IL-8 and a higher number of neutrophils were identified in samples from the H1N1 group when compared to the COVID-19 group. Discussion: It is suggested that the SARS-CoV-2 virus evokes an exacerbated response of the host's immune system but differs from that observed in the H1N1pdm09 disease because it may not be trigger by Th17 response. With the low expression of IL-8, IL-17A, neutrophil recruitment to the site of infection becomes impaired, resulting in viral persistence. On the other hand, in the COVID-19 disease, the immune response by Th2 cells seems to be exacerbated, observed by the extent of the lung injury. This uncontrolled response and, mainly, the lack of a therapeutic target, culminates in disease progression and, consequently, in shorter survival time.

Alveolar Neutrophilic Recruitment in COVID-19 May Not be Mediated by Th17 Response

Introduction: The COVID-19 current pandemic disease differs from the H1N1pdm09 caused by Influenza A virus H1N1 subtype, by how it induces a pro-inflammatory response in infected lungs. Objective: Investigate the role of Th17 response in the pathogenesis of COVID-19 injury by analyzing the tissue expression of interleukins 8 and 17A and the neutrophils in lung samples of patients who die of COVID-19, comparing to H1N1pdm09. Study design and post-mortem results: Six lung samples from patients SARS-CoV-2 infected (COVID-19 group), and ten lung samples from adults who died from H1N1pdm09 (H1N1 group), were tested. A control group was also added to the study. H&E slides were used for neutrophils scoring. The tissue expression of IL-8 and IL-17A was identified by immunohistochemistry. Tissue expression increasing of IL-17A and IL-8 and a higher number of neutrophils were identified in samples from the H1N1 group when compared to the COVID-19 group. Discussion: It is suggested that the SARS-CoV-2 virus evokes an exacerbated response of the host's immune system but differs from that observed in the H1N1pdm09 disease because it may not be trigger by Th17 response. With the low expression of IL-8, IL-17A, neutrophil recruitment to the site of infection becomes impaired, resulting in viral persistence. On the other hand, in the COVID-19 disease, the immune response by Th2 cells seems to be exacerbated, observed by the extent of the lung injury. This uncontrolled response and, mainly, the lack of a therapeutic target, culminates in disease progression and, consequently, in shorter survival time.

Identification of combinatorial and singular genomic signatures of host adaptation in influenza A H1N1 and H3N2 subtypes

BackgroundThe underlying strategies used by influenza A viruses (IAVs) to adapt to new hosts while crossing the species barrier are complex and yet to be understood completely. Several studies have been published identifying singular genomic signatures that indicate such a host switch. The complexity of the problem suggested that in addition to the singular signatures, there might be a combinatorial use of such genomic features, in nature, defining adaptation to hosts..ResultsWe used computational rule-based modeling to identify combinatorial sets of interacting amino acid (aa) residues in 12 proteins of IAVs of H1N1 and H3N2 subtypes. We built highly accurate rule-based models for each protein that could differentiate between viral aa sequences coming from avian and human hosts,. We found 68 combinations of aa residues associated to host adaptation (HAd) on HA, M1, M2, NP, NS1, NEP, PA, PA-X, PB1 and PB2 proteins of the H1N1 subtype and 24 on M1, M2, NEP, PB1 and PB2 proteins of the H3N2 subtypes. In addition to these combinations, we found 132 novel singular aa signatures distributed among all proteins, including the newly discovered PA-X protein, of both subtypes. We showed that HA, NA, NP, NS1, NEP, PA-X and PA proteins of the H1N1 subtype carry H1N1-specific and HA, NA, PA-X, PA, PB1-F2 and PB1 of the H3N2 subtype carry H3N2-specific HAd signatures. M1, M2, PB1-F2, PB1 and PB2 of H1N1 subtype, in addition to H1N1 signatures, also carry H3N2 signatures. Similarly M1, M2, NP, NS1, NEP and PB2 of H3N2 subtype were shown to carry both H3N2 and H1N1 HAd signatures.ConclusionsTo sum it up, we computationally constructed simple IF-THEN rule-based models that could distinguish between aa sequences of virus particles originating from avian and human hosts. From the rules we identified combinations of aa residues as signatures facilitating the adaptation to specific hosts. The identification of combinatorial aa signatures suggests that the process of adaptation of IAVs to a new host is more complex than previously suggested. The present study provides a basis for further detailed studies with the aim to elucidate the molecular mechanisms providing the foundation for the adaptation process.