Ceren Ciraci

Immune complexes (ICs) skew immune responses toward either a pro- or anti-inflammatory direction based on the type of stimulation. Immunoglobulin E (IgE) is associated with Th2 immune responses and known to activate innate immune cells. However, roles of antigen (Ag)-specific-IgE ICs in regulating human eosinophil responses remain elusive; therefore, this study builts upon the mechanism of which ovalbumin (Ova)-IgE ICs affects eosinophilic responses utilizing human EoL-1 cell line as a model. Eosinophils are granulocytes functioning through pattern recognition receptors (PRRs) and destructive granule contents in allergic inflammation and parasitic infections. One of the PRRs that eosinophils express is NLRC4, a member of the CARD domain containing nucleotide-binding oligomerization (NOD)-like receptor (NLR) family. Upon recognition of its specific ligand flagellin, NLRC4 inflammasome is formed and leads to the release of interleukin-1β (IL-1β). We exhibited that Ova-IgE ICs induced the NLRC4-inflammasome components, including NLRC4, caspase-1, intracellular IL-1β, and secretion of IL-1β, as well as the granule contents MMP9, TIMP1, and TIMP2 proteins via TLR2 signaling; these responses were suppressed, when NLRC4 inflammasome got actived in the presence of ICs. Furthermore, Ova-IgE ICs induced mRNA expressions of MMP9, TIMP2, and ECP and protein expressions of MMP9 and TIMP2 in EoL-1 through FcɛRII. Interestingly, TLR2 ligand and Ova-IgE ICs costimulation elevated the number of CD63 cells, a degranulation marker, as compared to the native IgE. Collectively, our findings provide a mechanism for the impacts of Ova-IgE ICs on eosinophilic responses via NLRC4-inflammasome and may help understand eosinophil-associated diseases, including chronic eosinophilic pneumonia, eosinophilic esophagitis, eosinophilic granulomatosis, parasitic infections, allergy, and asthma.

Eosinophils function in rapid innate immune responses and allergic reactions. Recent research has raised the possibility that the histone deacetylase inhibitor valproic acid (VPA) may be a promising therapeutic agent for treatment of allergic responses and certain cancers. However, its effects on eosinophils remain unclear. Utilizing the EoL-1 human eosinophil cell line as a model, we investigated the effects of VPA on oxidative stress- and autophagy-mediated immune responses. We found that VPA induced reactive oxidative species (ROS) generation and eosinophil activation without affecting cell viability. Moreover, VPA treatment suppressed the negative regulator of antioxidant transcription factor Nrf2, which is known to activate antioxidant defense. Interestingly, VPA was able to increase autophagic markers, as well as NLRP3 and NLRC4 mRNA activation, in Eol-1 cells in a dose-dependent manner. Collectively, our results indicate that VPA could increase the severity of allergic responses, and if so, it clearly would not be a suitable drug for the treatment of allergic reactions. However, VPA does have the potential to induce autophagy and to regulate the inflammatory responses via inflammasome-driven caspase-1 deactivation in a dose-dependent manner.

The advent of new technologies in gene expression, immunology, molecular biology, and computational modeling studies has expedited the discovery process and provided us with a holistic view of host immune responses that are highly regulated. The regulatory mechanisms of the immune system lie not only in weakening the attacker directly but also in fortifying the defender for the development of an efficient adaptive immune response. This chapter reviews a comprehensive set of experimental and bioinformatic studies designed to deepen the current knowledge on the regulatory T cells (Tregs) in the context of Pattern Recognition Receptors (PRRs). Initially, we examined both membrane-bound Toll-like Receptors (TLRs) and C Type Lectin Receptors (CLRs); and cytosolic NOD-like Receptors (NLRs) and RIG-I like Receptors (RLRs) in Tregs. Then, we revisited the disease conditions associated with regulatory T cells by emphasizing the essential roles of PRRs. Expanding our knowledge and strategies on the regulatory mechanisms are likely to provide our best chances for long-term disease control and maintenance of homeostasis.

The human angiotensin-converting enzyme 2 (hACE2) receptor is the primary receptor for SARS-CoV-2 infection. However, the presence of alternative receptors such as the transmembrane glycoprotein CD147 has been proposed as a potential route for SARS-CoV-2 infection. The outcomes of SARS-CoV-2 spike protein binding to receptors have been shown to vary among individuals. Additionally, some patients infected with SARS-CoV-2 develop autoimmune responses. Given that CD147 is involved in the hyperactivation of memory T cells resulting in autoimmunity, we investigated the interaction of the SARS-CoV-2 viral spike protein with CD147 receptor and retinal specific CD147 Ig0 domain in silico using molecular docking and molecular dynamics (MD) simulations. The results indicated that binding involves two critical residues Lys63 and Asp65 in a ubiquitous CD147 isoform, potentially leading to the hyperactivation of T cells for only SARS-CoV-2, but not for SARS-CoV or MERS-CoV. Overall binding was confirmed by docking simulations. Next, MD analyses were completed to verify the docking poses. Polar interactions suggested that the interaction via Lys63 and Asp65 might be one of the determinants associated with severe COVID-19 outcomes. Neither did SARS-CoV nor MERS-CoV bind to these two critical residues when molecular docking analyses were performed. Interestingly, SARS-CoV was able to bind to CD147 with a lower affinity (-4.5 kcal/mol) than SARS-CoV-2 (-5.6 kcal/mol). Furthermore, Delta and Omicron variants of SARS-CoV-2 did not affect the polar interactions with Lys63 and Asp65 in CD147. This study further strengthens the link between SARS-CoV-2 infection and autoimmune responses and provides novel insights for prudent antiviral drug designs for COVID-19 treatment that have implications in the prevention of T cell hyperactivation.

Akkaya I, et al. Immune Netw. 2021 Nov;21(6):e42. https://doi.org/10.4110/in.2021.21.e42

NLRP11 is a member of the PYD domain‐containing, nucleotide‐binding oligomerization‐domain (NOD‐) like receptor (NLR) family. The true stimulus of NLRP11 is still unclear to date, so the current study is built upon NLRP11 induction via adenosine stimulation and that activation can shape adaptive immune responses in a caspase‐1‐independent manner. We examined the regulation and mechanism of adenosine responsiveness via NLRP11 in human Daudi Burkitt’s B lymphoma cells and their effects on human peripheral CD4 ⁺ T lymphocytes from healthy individuals. NLRP11 was significantly upregulated after induction with adenosine at both the mRNA and protein levels, which led to the interaction of endogenous NLRP11 with the ASC adaptor protein; however, this interaction did not result in the activation of the caspase‐1 enzyme. Furthermore, cocultures of NLRP11‐expressing Burkitt’s lymphoma cells and naïve human peripheral CD4 ⁺ T lymphocytes had reduced IFN‐ γ and IL‐17A production, whereas IL‐13 and IL‐10 cytokines did not change. Interestingly, IFN‐ γ and IL‐17A were recovered after transfection of Burkitt’s lymphoma cells with siRNAs targeting NLRP11. Concomitant with NLRP11 upregulation, we also exhibited that adenosine A _2B receptor signaling induced two phosphorylated downstream effectors, pErk1/2 and pAkt (Ser473), but not pAkt (Thr308). Taken together, our data indicate that adenosine is a negative regulator of Th1 and Th17 responses via NLRP11 in an inflammasome‐independent manner.

Immune regulation is an essential feature of immune responses. The failure of such regulation results in allergic reactions and debilitating autoimmune diseases that can be fatal. Furthermore, the recent increase in the prevalence of the latter as well as the medical severity makes this a subject of great medical interest. Autoimmunity results from a breakdown in or the failure of the self-tolerance mechanisms. Many genes have been identified in which mutations cause the predisposition to autoinflammation and autoimmunity in human and in animal models. The relatively small number of genes explored to date unquestionably shows the challenges of identifying the associated genes in outbred populations of humans. One chief contributing gene family to both autoinflammatory and autoimmune diseases is the nucleotide-binding and oligomerization domain (NOD)-like receptor (NLR) family. Ever since their discovery, NLRs have drawn considerable attention for their ability to form multiprotein complexes called inflammasomes and also for their roles as NLRs, independent of inflammasome complexes. We herein first revisit general characteristics of NLRs and inflammasomes. We then couple this knowledge with the most recent findings related to autoinflammatory and autoimmune diseases, while highlighting some unanswered questions and future perspectives in elucidating NLR roles in health and disease.

Background: NLRC5 is a member of the CARD domain containing, nucleotide-binding oligomerization (NOD)-like receptor (NLR) family, which recognizes pathogen-associated molecular patterns (PAMPs) and initiates an innate immune response leading to inflammation and/or cell death. However, the specific role of NLRC5 as a modulator of the inflammatory immune response remains controversial. It has been reported to be a mediator of type I IFNs, NF-kB, and MHC class I gene. But no study on NLRC5 function has been reported to date in chickens. In the current study, we investigated the role of NLRC5 in the regulation of IFNA, IFNB, IL-6, and MHC class I in the chicken HD11 macrophage cell line, by using RNAi technology. HD11 cells were transfected with one of five siRNAs (s1, s2, s3, negative-siRNA, or a mixture of s1, s2, s3-siRNAs). After 24 hours, cells were exposed to LPS or poly (I:C) or a vehicle control. Gene expression of NLRC5, IFNA, IFNB, IL-6, and MHC class I at 2, 4, 6, and 8 hours post stimulation (hps) was quantified by qPCR. Results: The expression of NLRC5, IFNA, IFNB, and IL-6 genes in negative irrelevant transfection controls was up-regulated at 2 hps after LPS treatment compared to the vehicle controls. S3-siRNA effectively knocked down NLRC5 expression at 4 hps, and the expression of IFNA and IFNB (but not IL-6 and MHC class I) was also down-regulated at 4 hps in s3-siRNA transfected cells, compared to negative irrelevant transfection controls. Stimulation by LPS appeared to relatively restore the decrease in NLRC5, IFNA, and IFNB expression, but the difference is not significant. Conclusions: Functional characterization of chicken NLRC5 in an in vitro system demonstrated its importance in regulating intracellular molecules involved in inflammatory response. The knockdown of NLRC5 expression negatively mediates gene expression of IFNA and IFNB in the chicken HD11 cell line; therefore, NLRC5 likely has a role in positive regulation of IFNA and IFNB expression. No direct relationship was found between NLRC5 knockdown and IL-6 and MHC class I expression. Future studies will further clarify the roles of NLRC5 and other NLRs in infectious diseases of chickens and may increase the efficacy of antiviral vaccine design.