Prenatal Infection by Respiratory Viruses Is Associated with Immunoinflammatory Responses in the Fetus.

Rationale: Respiratory viral infections can be transmitted from pregnant women to their offspring, but frequency, mechanisms, and postnatal outcomes remain unclear. Objectives: The aims of this prospective cohort study were to compare the frequencies of transplacental transmission of respiratory syncytial virus (RSV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), analyze the concentrations of inflammatory mediators in maternal and fetal blood, and assess clinical consequences. Methods: We recruited pregnant women who developed upper respiratory infections or tested positive for SARS-CoV-2. Maternal and cord blood samples were collected at delivery. Study questionnaires and electronic medical records were used to document demographic and medical information. Measurements and Main Results: From October 2020 to June 2022, droplet digital PCR was used to test blood mononuclear cells from 103 mother-baby dyads. Twice more newborns in our sample were vertically infected with RSV compared with SARS-CoV-2 (25.2% [26 of 103] vs. 11.9% [12 of 101]; P = 0.019). Multiplex ELISA measured significantly increased concentrations of several inflammatory cytokines and chemokines in maternal and cord blood from newborns, with evidence of viral exposure in utero compared with control dyads. Prenatal infection was associated with significantly lower birth weight and postnatal weight growth. Conclusions: Data suggest a higher frequency of vertical transmission for RSV than SARS-CoV-2. Intrauterine exposure is associated with fetal inflammation driven by soluble inflammatory mediators, with expression profiles dependent on the virus type and affecting the rate of viral transmission. Virus-induced inflammation may have pathological consequences already in the first days of life, as shown by its effects on birth weight and postnatal weight growth.

Immune Response to Respiratory Viral Infections.

The respiratory system is constantly exposed to viral infections that are responsible for mild to severe diseases. In this narrative review, we focalized the attention on respiratory syncytial virus (RSV), influenza virus, and severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) infections, responsible for high morbidity and mortality in the last decades. We reviewed the human innate and adaptive immune responses in the airways following infection, focusing on a particular population: newborns and pregnant women. The recent Coronavirus disease-2019 (COVID-19) pandemic has highlighted how our interest in viral pathologies must not decrease. Furthermore, we must increase our knowledge of infection mechanisms to improve our future defense strategies.

Respiratory syncytial virus infects peripheral and spinal nerves and induces chemokine-mediated neuropathy.

Respiratory syncytial virus (RSV) primarily infects the respiratory epithelium, but growing evidence suggests it may also be responsible for neurological sequelae. In 3D microphysiological peripheral nerve cultures, RSV infected neurons, macrophages, and dendritic cells along two distinct trajectories depending on the initial viral load. Low-level infection was transient, primarily involved macrophages, and induced moderate chemokine release with transient neural hypersensitivity. Infection with higher viral loads was persistent, infected neuronal cells in addition to monocytes, and induced robust chemokine release followed by progressive neurotoxicity. In spinal cord cultures, RSV infected microglia and dendritic cells but not neurons, producing a moderate chemokine expression pattern. The persistence of infection was variable but could be identified in dendritic cells as long as 30 days post-inoculation. This study suggests that RSV can disrupt neuronal function directly through infection of peripheral neurons and indirectly through infection of resident monocytes, and inflammatory chemokines likely mediate both mechanisms.

RSV attenuates epithelial cell restitution by inhibiting actin cytoskeleton-dependent cell migration.

The airway epithelium's ability to repair itself after injury, known as epithelial restitution, is an essential mechanism enabling the respiratory tract's normal functions. Respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract infections worldwide. We sought to determine whether RSV delays the airway epithelium wound repair process both in vitro and in vivo. We found that RSV infection attenuated epithelial cell migration, a step in wound repair, promoted stress fiber formation, and mediated assembly of large focal adhesions. Inhibition of Rho-associated kinase, a master regulator of actin function, reversed these effects. There was increased RhoA and phospho-myosin light chain 2 following RSV infection. In vivo, mice were intraperitoneally inoculated with naphthalene to induce lung injury, followed by RSV infection. RSV infection delayed reepithelialization. There were increased concentrations of phospho-myosin light chain 2 in day 7 naphthalene + RSV animals, which normalized by day 14. This study suggests a key mechanism by which RSV infection delays wound healing.

RSV infection potentiates TRPV1-mediated calcium transport in bronchial epithelium of asthmatic children.

The transient receptor potential vanilloid 1 (TRPV1) channel is expressed in human bronchial epithelium (HBE), where it transduces Ca2+ in response to airborne irritants. TRPV1 activation results in bronchoconstriction, cough, and mucus production, and may therefore contribute to the pathophysiology of obstructive airway disease. Since children with asthma face the greatest risk of developing virus-induced airway obstruction, we hypothesized that changes in TRPV1 expression, localization, and function in the airway epithelium may play a role in bronchiolitis and asthma in childhood. We sought to measure TRPV1 protein expression, localization, and function in HBE cells from children with versus without asthma, both at baseline and after RSV infection. We determined changes in TRPV1 protein expression, subcellular localization, and function both at baseline and after RSV infection in primary HBE cells from normal children and children with asthma. Basal TRPV1 protein expression was higher in HBE from children with versus without asthma and primarily localized to plasma membranes (PMs). During RSV infection, TRPV1 protein increased more in the PM of asthmatic HBE as compared with nonasthmatic cells. TRPV1-mediated increase in intracellular Ca2+ was greater in RSV-infected asthmatic cells, but this increase was attenuated when extracellular Ca2+ was removed. Nerve growth factor (NGF) recapitulated the effect of RSV on TRPV1 activation in HBE cells. Our data suggest that children with asthma have intrinsically hyperreactive airways due in part to higher TRPV1-mediated Ca2+ influx across epithelial membranes, and this abnormality is further exacerbated by NGF overexpression during RSV infection driving additional Ca2+ from intracellular stores.

Titanium dioxide nanoparticles exaggerate respiratory syncytial virus-induced airway epithelial barrier dysfunction.

Respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract infections in children worldwide. While most develop a mild, self-limiting illness, some develop severe acute lower respiratory infection and persistent airway disease. Exposure to ambient particulate matter has been linked to asthma, bronchitis, and viral infection in multiple epidemiological studies. We hypothesized that coexposure to nanoparticles worsens RSV-induced airway epithelial barrier dysfunction. Bronchial epithelial cells were incubated with titanium dioxide nanoparticles (TiO2-NP) or a combination of TiO2-NP and RSV. Structure and function of epithelial cell barrier were analyzed. Viral titer and the role of reactive oxygen species (ROS) generation were evaluated. In vivo, mice were intranasally incubated with TiO2-NP, RSV, or a combination. Lungs and bronchoalveolar lavage (BAL) fluid were harvested for analysis of airway inflammation and apical junctional complex (AJC) disruption. RSV-induced AJC disruption was amplified by TiO2-NP. Nanoparticle exposure increased viral infection in epithelial cells. TiO2-NP induced generation of ROS, and pretreatment with antioxidant, N-acetylcysteine, reversed said barrier dysfunction. In vivo, RSV-induced injury and AJC disruption were augmented in the lungs of mice given TiO2-NP. Airway inflammation was exacerbated, as evidenced by increased white blood cell infiltration into the BAL, along with exaggeration of peribronchial inflammation and AJC disruption. These data demonstrate that TiO2-NP exposure exacerbates RSV-induced AJC dysfunction and increases inflammation by mechanisms involving generation of ROS. Further studies are required to determine whether NP exposure plays a role in the health disparities of asthma and other lung diseases, and why some children experience more severe airway disease with RSV infection.

Effects of maternal-fetal transmission of viruses and other environmental agents on lung development.

New information is emerging concerning the influence of environmental factors (e.g., viruses, pollutants, nutrients) on fetal lung development and the prenatal modulation of cellular and molecular effectors essential to the control of airway function, which may shed new light into the pathogenesis of chronic obstructive pulmonary disease in childhood. In particular, recent studies have shown that nanosize biological and inorganic particles (e.g., respiratory viruses and pollutants) are able to spread hematogenously across the placenta from mother to offspring and interfere with lung development during critical "windows of opportunity". Furthermore, the nutritional balance of maternal diet during pregnancy can affect postnatal lung structure and function. Adverse prenatal environmental conditions can predispose to increased airway reactivity by inducing aberrant cholinergic innervation of the respiratory tract, enhanced contractility of the airway smooth muscle, and impaired innate immunity. Such changes can persist long after birth and might provide a plausible explanation to the development of chronic airway dysfunction in children, even in the absence of atopic predisposition. Insight into maternal-fetal interactions will contribute to a better understanding of the pathogenesis of highly prevalent diseases like bronchiolitis and asthma, and may lead to more precise preventative and therapeutic strategies, or new indications for existing ones.

Disruption of the airway epithelial barrier in a murine model of respiratory syncytial virus infection.

Respiratory syncytial virus (RSV) is a major cause of hospitalization for infants and young children worldwide. RSV is known to infect epithelial cells and increase the permeability of model airway epithelial monolayers in vitro. We hypothesized that RSV infection also induces airway barrier dysfunction in vivo. C57BL/6 mice were intranasally inoculated with RSV, and on day 4 post-inoculation were examined for viral replication, lung inflammation, and barrier integrity as well as the structure and molecular composition of epithelial junctions. In parallel, primary mouse tracheal epithelial cells (mTEC) were cultured for in vitro studies. RSV-infected mice lost weight and showed significant peribronchial inflammation compared with noninfected controls and UV-inactivated RSV-inoculated animals. RSV infection increased the permeability of the airway epithelial barrier and altered the molecular composition of epithelial tight junctions. The observed RSV-induced barrier disruption was accompanied by decreased expression of several tight-junction proteins and accumulation of cleaved extracellular fragments of E-cadherin in bronchoalveolar lavage and mTEC supernatants. Similarly, in vitro RSV infection of mTEC monolayers resulted in enhanced permeability and disruption of tight-junction structure. Furthermore, incubation of mTEC monolayers with a recombinant fragment of E-cadherin caused tight-junction disassembly. Taken together, these data indicate that RSV infection leads to airway barrier dysfunction in vivo, mediated by either decreased expression or cleavage of junctional proteins. Our observations provide further insights into the pathophysiology of RSV infection and provide a rationale for development of barrier-protecting agents to alleviate the pathogenic effects of RSV infection.

Induction of high-mobility group Box-1 in vitro and in vivo by respiratory syncytial virus.

BackgroundDespite decades that have passed since its discovery, accurate biomarkers of respiratory syncytial virus (RSV) disease activity and effective therapeutic strategies are still lacking. The high-mobility group box type 1 (HMGB1) protein has been proposed as a possible link between RSV and immune system, but only limited information is currently available to support this hypothesis.MethodsExpression of HMGB1 gene and protein was analyzed by quantitative PCR, enzyme-linked immunosorbent assay (ELISA), western blot, immunocytochemistry, and confocal microscopy in immortalized and primary human bronchial epithelial cells, as well as in rat pup lungs. The role of HMGB1 in RSV infection was explored using glycyrrhizin, a selective HMGB1 inhibitor.ResultsRSV infection strongly induced HMGB1 expression both in vitro and in vivo. Glycyrrhizin dose-dependently inhibited HMGB1 upregulation in both RSV-infected immortalized and primary human bronchial epithelial cells, and this effect was associated with significant reduction of viral replication.ConclusionOur data suggest that HMGB1 expression increases during RSV replication. This seems to have a critical pathogenic role as its selective inhibition virtually modified the infection. These observations provide further insight into the pathophysiology of RSV infection and uncover a potential biomarker and therapeutic target for the most common respiratory infection of infancy.

Nanoparticles-induced apoptosis of human airway epithelium is mediated by proNGF/p75NTR signaling.

Environmental and occupational exposures to respirable ultrafine fractions of particulate matter (PM) have been implicated in the initiation and exacerbation of lung diseases. However, the precise mechanisms underlying production of cell damage and death attributed to nanoparticles (NP) on human airway epithelium are not fully understood. This study examined the role of neurotrophic pathways in NP-induced airway toxicity. Size and agglomeration of TiO2 nanoparticles (TiO2-NP) and fine (TiO2-FP) particles were measured by dynamic light scattering. Expression and signaling of key neurotrophic factors and receptors were assessed by real-time polymerase chain reaction, flow cytometry, immunostaining, and Western blot in various respiratory epithelial cells after exposure to TiO2-NP or TiO2-FP. Particle-induced cell death was measured by flow cytometry after annexin V/propidium iodide staining. The role of neurotrophin-dependent apoptotic pathways was analyzed with specific blocking antibodies or siRNAs. Exposure of human epithelial cells to TiO2-NP enhanced interleukin (IL)-1α synthesis, as well as nerve growth factor (NGF) gene expression and protein levels, specifically the precursor form (proNGF). TiO2-NP exposure also increased expression of p75NRF receptor genes. These neurotropic factor and receptor responses were stimulated by IL-1α and abolished by its specific receptor antagonist (IL-1-ra). TiO2-NP also increased JNK phosphorylation and apoptosis, which was prevented by anti-p75NRF or NGFsiRNA. Data demonstrated that TiO2-NP exerted adverse effects in the respiratory tract by inducing unbalanced overexpression of immature neurotrophins, which led to apoptotic death of epithelial cells signaled through the death receptor p75NTR. This may result in airway inflammation and hyperreactivity after exposure to TiO2-NP.

Maternal high-fat hypercaloric diet during pregnancy results in persistent metabolic and respiratory abnormalities in offspring.

BACKGROUND: We have shown in a previous population-based study significant correlation between childhood asthma and early abnormalities of lipid and glucose metabolism. This study's specific aim was to determine whether maternal nutrition in pregnancy affects postnatal metabolic and respiratory outcomes in the offspring. METHODS: On gestation day 1, dams were switched from standard chow to either high-fat hypercaloric diet or control diet. Terminal experiments were performed on newborn and weanling offspring of dams fed the study diet during gestation and lactation, and on adult offspring maintained on the same diet as their mother. RESULTS: Pups born from high-fat hypercaloric diet (HFD) dams developed metabolic abnormalities persistent throughout development. Cytokine expression analysis of lung tissues from newborns born to HFD dams revealed a strong proinflammatory pattern. Gene expression of neurotrophic factors and receptors was upregulated in lungs of weanlings born to HFD dams, and this was associated to higher respiratory system resistance and lower compliance at baseline, as well as hyperreactivity to aerosolized methacholine. Furthermore, HFD dams delivered pups prone to develop more severe disease after respiratory syncytial virus (RSV) infection. CONCLUSION: Maternal nutrition in pregnancy is a critical determinant of airway inflammation and hyperreactivity in offspring and also increases risk for bronchiolitis independent from prepregnancy nutrition.

Elevated IL-3 and IL-12p40 levels in the lower airway of infants with RSV-induced bronchiolitis correlate with recurrent wheezing.

Respiratory Syncytial Virus (RSV) is the first cause of hospitalization due to bronchiolitis in infants. RSV bronchiolitis has been linked to asthma and recurrent wheezing, however the mechanisms behind this association have not been elucidated. Here, we evaluated the cytokine and chemokine profiles in the airways in infants with RSV bronchiolitis. Nasopharyngeal Aspirates (NPA) and Bronchoalveolar Lavage Fluids (BALF) from infants hospitalized due to RSV bronchiolitis and healthy controls were analyzed for cytokine and chemokine production. We observed elevated levels of Th2 cytokines (IL-3, IL-4, IL-10 and IL-13), pro-inflammatory cytokines and chemokines (IL-1ß, IL-6, TNF-ß, MCP-1/CCL2, MIP-1α/CCL3 and IL-8/CXCL8) in BALF from infants with RSV bronchiolitis, as compared to controls. We found a direct correlation of IL-3 and IL-12p40 levels with the development of recurrent wheezing later in life. These results suggest that IL-3 and IL-12p40 could be considered as molecular predictors for recurrent wheezing due to RSV infection.

Sinus and adenoid inflammation in children with chronic rhinosinusitis and asthma.

BACKGROUND: Chronic rhinosinusitis (CRS) and asthma frequently coexist in children and adults. However, the precise pathophysiologic mechanism of this interaction is still poorly understood, especially in children, owing to the lack of direct measurements of mucosal inflammation in the upper airways. OBJECTIVE: To determine the pathophysiologic mechanism by analyzing the expression of a large array of inflammatory cytokines and chemokines in the sinus and adenoid tissues surgically removed from pediatric patients with CRS refractory to medical management. METHODS: Twenty-eight children 2 to 12 years old diagnosed with CRS with or without asthma and 10 controls were included in this prospective, nonrandomized study. Mucosal expression of 40 inflammatory cytokines was measured with a multiplex assay and was normalized to total tissue protein. RESULTS: Compared with children with CRS and without asthma, children with CRS and asthma had significantly higher sinus levels of tumor necrosis factor-α and adenoid levels of epidermal growth factor, eotaxin, fibroblast growth factor-2, growth-related oncogene, and platelet-derived growth factor-AA. CONCLUSION: The inflammatory response in the upper airway mucosa of children with asthma and CRS was similar, but more severe, compared with children with CRS without asthma. This observation is consistent with the hypothesis that asthma in these patients is caused or exacerbated by severe upper airway disease and supports the concept that treating sinus disease is paramount in the management of chronic asthma in children using, for the first time, direct measurements of airway inflammation in children.

Biomarkers of respiratory syncytial virus (RSV) infection: specific neutrophil and cytokine levels provide increased accuracy in predicting disease severity.

Despite fundamental advances in the research on respiratory syncytial virus (RSV) since its initial identification almost 60 years ago, recurring failures in developing vaccines and pharmacologic strategies effective in controlling the infection have allowed RSV to become a leading cause of global infant morbidity and mortality. Indeed, the burden of this infection on families and health care organizations worldwide continues to escalate and its financial costs are growing. Furthermore, strong epidemiologic evidence indicates that early-life lower respiratory tract infections caused by RSV lead to the development of recurrent wheezing and childhood asthma. While some progress has been made in the identification of reliable biomarkers for RSV bronchiolitis, a "one size fits all" biomarker capable of accurately and consistently predicting disease severity and post-acute outcomes has yet to be discovered. Therefore, it is of great importance on a global scale to identify useful biomarkers for this infection that will allow pediatricians to cost-effectively predict the clinical course of the disease, as well as monitor the efficacy of new therapeutic strategies.

Passive antibody transfer from pregnant women to their fetus are maximized after SARS-CoV-2 vaccination irrespective of prior infection.

Background: Pregnancy is associated with a higher risk of adverse symptoms and outcomes for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection for both mother and neonate. Antibodies can provide protection against SARS-CoV-2 infection and are induced in pregnant women after vaccination or infection. Passive transfer of these antibodies from mother to fetus in utero may provide protection to the neonate against infection. However, it is unclear whether the magnitude or quality and kinetics of maternally derived fetal antibodies differs in the context of maternal infection or vaccination. Objective: We aimed to determine whether antibodies transferred from maternal to fetus differed in quality or quantity between infection- or vaccination-induced humoral immune responses. Methods: We evaluated 93 paired maternal and neonatal umbilical cord blood plasma samples collected between October 2020 and February 2022 from a birth cohort of pregnant women from New Orleans, Louisiana, with histories of SARS-CoV-2 infection and/or vaccination. Plasma was profiled for the levels of spike-specific antibodies and induction of antiviral humoral immune functions, including neutralization and Fc-mediated innate immune effector functions. Responses were compared between 4 groups according to maternal infection and vaccination. Results: We found that SARS-CoV-2 vaccination or infection during pregnancy increased the levels of antiviral antibodies compared to naive subjects. Vaccinated mothers and cord samples had the highest anti-spike antibody levels and antiviral function independent of the time of vaccination during pregnancy. Conclusions: These results show that the most effective passive transfer of functional antibodies against SARS-CoV-2 in utero is achieved through vaccination, highlighting the importance of vaccination in pregnant women.

Respiratory syncytial virus and asthma: speed-dating or long-term relationship?

PURPOSE OF REVIEW: Respiratory syncytial virus (RSV) is the most common respiratory pathogen in infants and young children worldwide. Furthermore, epidemiological evidence has been accumulating that RSV lower respiratory tract infection in infants may be linked to subsequent development of recurrent wheezing and asthma in childhood. This article reviews the epidemiological evidence linking RSV and asthma and some new hypotheses of the cellular and molecular mechanisms of postviral airway inflammation and hyperreactivity that have been proposed to explain the epidemiological link. RECENT FINDINGS: New epidemiological studies have suggested that viral pathogens other than RSV, especially human rhinoviruses (HRV), may play an important role in the inception of atopic asthma. Also, recent experimental evidence is challenging the widely accepted axiom that RSV is cleared from immunocompetent hosts within weeks from the onset of the infection. In particular, bone marrow stromal cells may be a frequent target of human RSV infection, develop structural and functional changes when infected, participate actively in the pathogenesis of the acute disease, and harbor the virus chronically, allowing persistence of the infection. SUMMARY: RSV - and possibly other common respiratory pathogens - play an important role not only in the exacerbation, but also in the inception of asthma. The latter effect may involve the persistence of latent virus in extrapulmonary tissues, similar to what has been recently found for some bacterial species. The most immediate consequence of these discoveries is that future prophylactic and therapeutic strategies for common infections caused by viral or bacterial pathogens may have to address the coverage of remote sites of latent persistence or replication, in order to avoid chronic sequelae-recurrent wheezing and asthma.

Impact of respiratory viral infections during pregnancy on the neurological outcomes of the newborn: current knowledge.

Brain development is a complex process that begins during pregnancy, and the events occurring during this sensitive period can affect the offspring's neurodevelopmental outcomes. Respiratory viral infections are frequently reported in pregnant women, and, in the last few decades, they have been related to numerous neuropsychiatric sequelae. Respiratory viruses can disrupt brain development by directly invading the fetal circulation through vertical transmission or inducing neuroinflammation through the maternal immune activation and production of inflammatory cytokines. Influenza virus gestational infection has been consistently associated with psychotic disorders, such as schizophrenia and autism spectrum disorder, while the recent pandemic raised some concerns regarding the effects of severe acute respiratory syndrome coronavirus 2 on neurodevelopmental outcomes of children born to affected mothers. In addition, emerging evidence supports the possible role of respiratory syncytial virus infection as a risk factor for adverse neuropsychiatric consequences. Understanding the mechanisms underlying developmental dysfunction allows for improving preventive strategies, early diagnosis, and prompt interventions.