Functional importance of bile acid-FXR signaling in neonatal immunity and disease

The neonatal period is a critical phase for the development of the intestinal immune system, marked by rapid adaptation to the external environment and unique nutritional demands. Breast milk plays a pivotal role in this transition, yet the mechanisms by which it influences neonatal mucosal immunity remain unclear. This review examines the potential mechanisms by which cell-free DNA (cfDNA) in breast milk may impact neonatal immune development, particularly through Toll-like receptor 9 (TLR9) signalling and gut microbiota interactions. We propose that cfDNA in breast milk interacts with TLR9 on the apical surface of neonatal intestinal epithelial cells, potentially serving as an initial anti-inflammatory stimulus before the establishment of commensal bacteria. This hypothesis is supported by the high concentration and stability of cfDNA in breast milk, as well as the known activation of TLR9 by mitochondrial DNA in breast milk. The review emphasises the need for further empirical research to validate these interactions and their implications for neonatal health, suggesting that understanding these dynamics could lead to improved strategies for neonatal care and disease prevention.

Chapter four - Immunoglobulins and their relevance in human milk

The multifaceted roles of breast milk antibodies

Cost-effectiveness of maternal immunization against neonatal invasive Group B Streptococcus in the Netherlands

Staphylococcus epidermidis accounts for the majority of cases of neonatal sepsis. Moreover, it has been demonstrated to be associated with neonatal morbidities, such as bronchopulmonary dysplasia (BPD), white matter injury (WMI), necrotizing enterocolitis (NEC) and retinopathy of prematurity (ROP), which affect short-term and long-term neonatal outcome. Imbalanced inflammation has been considered to be a major underlying mechanism of each entity. Conventionally regarded as a harmless commensal on human skin, S. epidermidis has received less attention than its more virulent relative Staphylococcus aureus. Particularities of neonatal innate immunity and nosocomial environmental factors, however, may contribute to the emergence of S. epidermidis as a significant nosocomial pathogen. Neonatal host response to S. epidermidis sepsis has not been fully elucidated. Evidence is emerging regarding the implication of S. epidermidis sepsis in the pathogenesis of neonatal inflammatory diseases. This review focuses on the interplay among S. epidermidis, neonatal innate immunity and inflammation-driven organ injury.

This review is a short synopsis of the roles cytokines play during fetal life, initiation of labor, and in neonatal immunity and diseases. Hematopoietic growth factors regulate the maturation of progenitors in fetal and neonatal hematopoietic organs. Cytokines act as extra-hematopoietic growth factors, modulators of feto-maternal tolerance and are involved in selective apoptosis during tissue remodeling. Inter-regulation of cytokine networks is critical for normal function and maturation of neonatal host defenses. Antigen specific immunity develops later in life and neonates initially depend on natural (innate) immunity. Cytokines regulate innate immunity and connect it with antigen specific adaptive immunity. Some cytokines have already found a place in routine NICU therapy (EPO and G-CSF), while diagnostic and therapeutic uses of others are under investigation (TPO, TNF-alpha, etc.).

Our environment poses a constant threat to our health. To survive, all organisms must be able to discriminate between good (food ingredients and microbes that help digest our food) and bad (pathogenic microbes, viruses and toxins). In vertebrates, discrimination between beneficial and harmful antigens mainly occurs at the mucosal surfaces of the respiratory, digestive, urinary and genital tract. Here, an extensive network of cells and organs form the basis of what we have come to know as the mucosal immune system. The mucosal immune system is composed of a single epithelial cell layer protected by a mucus layer. Different immune cells monitor the baso-lateral side of the epithelial cells and dispersed secondary lymphoid organs, such as Peyer’s patches and isolated lymphoid follicles are equipped with immune cells able to mount appropriate and specific responses. This review will focus on the current knowledge on host, dietary and bacterial-derived factors that shape the mucosal immune system before and after birth. We will discuss current knowledge on fetal immunity (both responsiveness and lymphoid organ development) as well as the impact of diet and microbial colonization on neonatal immunity and disease susceptibility. Lastly, inflammatory bowel disease will be discussed as an example of how the composition of the microbiota might predispose to disease later in life. A fundamental understanding of the mechanisms involved in mucosal immune development and tolerance will aid nutritional intervention strategies to improve health in neonatal and adult life.

Chronic immune-mediated diseases of adulthood often originate in early childhood. To investigate genetic associations between neonatal immunity and disease, we map expression quantitative trait loci (eQTLs) in resting myeloid cells and CD4+ T cells from cord blood samples, as well as in response to lipopolysaccharide (LPS) or phytohemagglutinin (PHA) stimulation, respectively. Cis-eQTLs are largely specific to cell type or stimulation, and 31% and 52% of genes with cis-eQTLs have response eQTLs (reQTLs) in myeloid cells and T cells, respectively. We identified cis regulatory factors acting as mediators of trans effects. There is extensive colocalisation between condition-specific neonatal cis-eQTLs and variants associated with immune-mediated diseases, in particular CTSH had widespread colocalisation across diseases. Mendelian randomisation shows causal neonatal gene expression effects on disease risk for BTN3A2, HLA-C and others. Our study elucidates the genetics of gene expression in neonatal immune cells, and aetiological origins of autoimmune and allergic diseases.

The mammalian Siglec receptor sialoadhesin (Siglec1, CD169) confers innate immunity against the encapsulated pathogen group B Streptococcus (GBS). Newborn lung macrophages have lower expression levels of sialoadhesin at birth compared with the postnatal period, increasing their susceptibility to GBS infection. In this study, we investigate the mechanisms regulating sialoadhesin expression in the newborn mouse lung. In both neonatal and adult mice, GBS lung infection reduced Siglec1 expression, potentially delaying acquisition of immunity in neonates. Suppression of Siglec1 expression required interactions between sialic acid on the GBS capsule and the inhibitory host receptor Siglec-E. The Siglec1 gene contains multiple STAT binding motifs, which could regulate expression of sialoadhesin downstream of innate immune signals. Although GBS infection reduced STAT1 expression in the lungs of wild-type newborn mice, we observed increased numbers of STAT1+ cells in Siglece−/− lungs. To test if innate immune activation could increase sialoadhesin at birth, we first demonstrated that treatment of neonatal lung macrophages ex vivo with inflammatory activators increased sialoadhesin expression. However, overcoming the low sialoadhesin expression at birth using in vivo prenatal exposures or treatments with inflammatory stimuli were not successful. The suppression of sialoadhesin expression by GBS–Siglec-E engagement may therefore contribute to disease pathogenesis in newborns and represent a challenging but potentially appealing therapeutic opportunity to augment immunity at birth.

Group B streptococci remain a serious cause of morbidity and mortality in neonates. GBS vaccine or immunoglobulin administered iv may enhance neonatal GBS immunity. Likewise, intrapartum antibiotic therapy of colonized mothers appears to reduce vertical transmission of group B streptococci and to prevent both maternal and neonatal GBS disease. However, the safety and effectiveness of routine penicillin prophylaxis less than or equal to 1 hr after birth remain in question. For example, penicillin prophylaxis appears to be of little value in infants with low birth weights (less than 2,000 g) who become symptomatic shortly after birth; however, it may reduce the incidence of disease in larger, full-term infants who acquire the group B streptococci at delivery or in the few hours immediately thereafter. The potential harm of administering penicillin to all neonates must also be considered, since routine antibiotic therapy may alter the incidence of both neonatal infections due to penicillin-resistant pathogens and possible later penicillin allergy. Theoretically, a single injection of penicillin at birth may suppress GBS disease in some neonates but not effectively treat it, allowing the disease to progress prior to diagnosis and therapy. The decision to use penicillin routinely in neonates to prevent GBS disease must therefore be made with caution. Presently, this decision must be made on a situational basis, with institutions having a high incidence of early-onset GBS disease electing to use penicillin only if the potential benefits outweigh the risks.

Studies have shown that IFNs have potent immunologic (1,2) and antiviral (3,4) activity in both in vitro and in vivo testing in the bovine species. In calves given IFN orally, enteric disease due to rotavirus or mixed rotavirus and coronavirus infection was prevented or reduced(5). Interferons are also involved in immunity to protozoal infections (6), and a polypeptide with many characteristics similar to an alpha IFN has been identified that stimulated macrophages to inhibit intracellular development of Eimeria bovis (7). In preliminary studies, calves administered partially purified natural bovine alpha IFN intravenously had less coccidial diarrhea than control calves (8).
 Although lymphocytes isolated from human colostrum and milk are capable of producing interferon ( IFN) when stimulated in vitro (9,10), assays of human colostrum and milk for IFN have usually yielded negative results (9-11). Since bovine peripheral blood lymphocytes, when stimulated with virus, are capable of producing alpha IFN (12), it would seem logical that these cells in mammary secretions would also have this ability. The only citation of the spontaneous occurrence of IFN in milk or colostrum from any species is a brief mention that a sample from one woman's breast milk contained antiviral activity (11). It was not known whether this woman was infected with a virus (11). It has been well documented, however, that when mother mice were inoculated with Newcastle disease virus, their milk contained measurable lFN (7). Furthermore, the newborns suckling such mothers had a significantly greater survival rate than that of controls after lethal challenge with vesicular stomatitis virus (7).
 Since most mammalian infants have a lower incidence of neonatal disease when raised on maternal milk as opposed to milk replacer (13,14), it was hypothesized that interferons in mammary secretions may play an important role in neonatal immunity. Since bovine milk and colostrum had not been heretofore examined, the objective of this study was to determine whether IFN could be found in detectable levels in normal bovine colostrum and milk.

Apoptotic inhibition and immune evasion have particular importance to efficient viral infection, while a dilemma often faced by viruses is that inhibiting apoptosis can up-regulate antiviral immune signaling. Herein, we uncovered that in addition to inhibiting caspase-8/extrinsic apoptosis, human cytomegalovirus (HCMV)-encoded UL36 suppresses interferon regulatory factor 3 (IRF3)-dependent immune signaling by directly targeting IRF3 to abrogate IRF3 interaction with stimulator of interferon genes or TANK-binding kinase 1 and inhibit IRF3 phosphorylation/activation. Although UL36-mediated caspase-8/extrinsic apoptosis inhibition enhances immune signaling, the immunosuppressing activity of UL36 counterbalances this immunoenhancing "side effect" undesirable for virus. Furthermore, we used mutational analyses to show that only the wild-type, but not the UL36 mutant losing either inhibitory activity, is sufficient to support effective HCMV replication in cells, showing the functional importance of the dual inhibition by UL36 for the HCMV life cycle. Together, our findings demonstrate a sophisticated mechanism by which HCMV tightly controls innate immune signaling and extrinsic apoptosis for efficient infection.

The immune system functions as a dynamic and sophisticated network that ensures efficient response to foreign antigens and tolerance to self-tissues.1 The function of the immune system relies on signal transduction that connects immune cells to the extracellular environment and mediates communication among the different types of immune cells. Among the well-characterized immunoregulatory signaling pathways is that which leads to the activation of nuclear factor-kappaB (NF-κB), a family of transcription factors that participates in different aspects of innate and adaptive immune responses.2, 3 NF-κB proteins normally exist as latent cytoplasmic complexes, but they can be rapidly activated in response to signals elicited from diverse immune receptors, including the pattern-recognition receptors on innate immune cells and the antigen receptors on lymphocytes.4, 5 Upon activation, NF-κB moves to the nucleus and participates in the induction of numerous genes, including those encoding pro-inflammatory cytokines, chemokines and cell adhesion molecules, which are important for the establishment of inflammation during the early phase of an infection. Although inflammation is a crucial immune mechanism against infections, inappropriately controlled inflammation contributes to the pathogenesis of immunological diseases. Thus, the NF-κB signaling pathway is subject to tight control by both positive and negative regulatory mechanisms.6 An important mechanism of NF-κB regulation is protein ubiquitination,7 which, like protein phosphorylation, has emerged as one of the most fundamental mechanisms of signal transduction.8 Originally linked to proteasomal degradation, ubiquitination is now being appreciated as a signal transduction mechanism that mediates both degradative and non-degradative cellular processes.9, 10 The complexity and specificity of ubiquitination are emphasized by the identification of more than 600 potential E3s in the human genome, which surpasses the number of protein kinases.11 Like phosphorylation, ubiquitination is a reversible process with the reverse reaction being catalyzed by the family of deubiquitinases or deubiquitinating enzymes (DUBs).12 An increasing number of E3s and DUBs have been assigned to the NF-κB signaling pathways, although only some of them have been characterized in depth by genetic approaches. There is no doubt that E3s and DUBs represent attractive candidates to be exploited as new therapeutic targets in the treatment of immunological disorders and cancer. This special issue of Cellular and Molecular Immunology presents four expert reviews that discuss the recent progress regarding innate immune receptor signaling, focusing on ubiquitination and NF-κB activation. Kingeter and Lin will discuss NF-κB activation by the C-type lectin receptors (CLRs), an emerging family of pattern-recognition receptors that recognize microbial carbohydrates and transduce signals via immunoreceptor tyrosine-based activation motifs. Analogous to the antigen receptors, the CLRs activate the IκB kinase by inducing IκB kinase ubiquitination via the CARD9–BCL10–MALT1 complex as well as IκB kinase phosphorylation through an unknown kinase. Since the CLR family includes many members whose ligands remain to be defined, it is anticipated that future studies will lead to important findings regarding the functions of CLRs. Two reviews focus on the mechanisms by which ubiquitination regulates NF-κB activation and immune receptor signaling. Jin and colleagues will discuss a newly characterized family of E3s, termed Peli (or Pellino), which mediates Toll-like receptor signaling in innate immune cells as well as regulates T-cell tolerance in the adaptive immune system. Shembade and Harhaj will focus on A20, a DUB with pivotal roles in the regulation of NF-κB signaling and inflammation. The recent linkage of A20 with human inflammatory and autoimmune diseases, as well as lymphoid malignancies, emphasizes the important function of A20 in the regulation of immune receptor signaling. Finally, Li and colleagues will present us with an example of how tight control of NF-κB and related signaling pathways ensures proper regulation of inflammation in respiratory infections. We sincerely hope that these reviews will offer the readers both the background knowledge and the current information in this rapidly developing area of Immunology research.

Mitochondria are well known as organelles responsible for the maintenance of cellular bioenergetics through the production of ATP. Although oxidative phosphorylation may be their most important function, mitochondria are also integral for the synthesis of metabolic precursors, calcium regulation, the production of reactive oxygen species, immune signaling, and apoptosis. Considering the breadth of their responsibilities, mitochondria are fundamental for cellular metabolism and homeostasis. Appreciating this significance, translational medicine has begun to investigate how mitochondrial dysfunction can represent a harbinger of disease. In this review, we provide a detailed overview of mitochondrial metabolism, cellular bioenergetics, mitochondrial dynamics, autophagy, mitochondrial damage-associated molecular patterns, mitochondria-mediated cell death pathways, and how mitochondrial dysfunction at any of these levels is associated with disease pathogenesis. Mitochondria-dependent pathways may thereby represent an attractive therapeutic target for ameliorating human disease.

Protein kinases are key components of multiple cell signaling pathways. Several receptor-like cytoplasmic kinases (RLCKs) have demonstrated roles in immune and developmental signaling across various plant species, making them of interest in the study of phosphorylation-based signal relay. Here, we present our investigation of a subgroup of RLCKs in Arabidopsis thaliana. Specifically, we focus on subgroup VIII RLCKs: MAZ and its paralog CARK6, as well as CARK7 and its paralog CARK9. We found that both MAZ and CARK7 associate with the calcium-dependent protein kinase CPK28 in planta and, furthermore, that CPK28 phosphorylates both MAZ and CARK7 on multiple residues in areas that are known to be critical for protein kinase activation. Genetic analysis suggested redundant roles for MAZ and CARK6 as negative regulators of the immune-triggered oxidative burst. We provide evidence that supports homo- and heterodimerization between CARK7 and MAZ, which may be a general feature of this subgroup. Multiple biochemical experiments indicated that neither MAZ nor CARK7 demonstrate catalytic protein kinase activity in vitro. Interestingly, we found that a mutant variant of MAZ incapable of protein kinase activity can complement maz-1 mutants, suggesting non-catalytic roles of MAZ in planta. Overall, our study identifies subgroup VIII RLCKs as new players in Arabidopsis immune signaling and highlights the importance of non-catalytic functions of protein kinases.