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HIV-1 assembly is initiated by the binding of Gag polyproteins to the inner leaflet of the plasma membrane, mediated by the myristylated matrix (MA) domain of Gag. Subsequent to membrane binding, Gag oligomerizes and buds as an immature, non-infectious virus particle, which, upon cleavage of the Gag precursor by the viral protease, transforms into a mature, infectious virion. During maturation, the MA lattice underlying the viral membrane undergoes a structural rearrangement and the newly released capsid (CA) protein forms a mature capsid that encloses the viral genome. While it is well established that formation of the mature capsid is essential to particle infectivity, the functional role of MA structural maturation remains unclear. Here, we examine MA maturation of an MA triple mutant, L20K/E73K/A82T, which exhibits distinct biochemical behaviours. The L20K/E73K/A82T mutant is a revertant derived by propagating the L20K mutant, which exhibits reduced infectivity and increased association of the Gag polyprotein with membranes. L20K/E73K/A82T replicates similarly to wild type but retains the increased Gag membrane binding properties of L20K. L20K/E73K/A82T MA also sediments to high-density fractions in sucrose gradients after detergent treatment under conditions that fully solubilize WT MA, suggesting enhanced MA-MA interactions. Cryo-electron tomography with subtomogram averaging reveals that the immature MA lattice of L20K/E73K/A82T closely resembles the wild type. However, mature virions of the triple mutant lack a detectable MA lattice, in stark contrast to both the wild type and L20K mutant. All-atom molecular dynamics simulations suggest that this absence results from destabilized inter-trimer interactions in the mature L20K/E73K/A82T MA. Furthermore, introducing additional mutations designed to disrupt the mature MA lattice does not impair particle infectivity. These findings suggest that an ordered, membrane-associated mature MA lattice is not essential for HIV-1 infectivity, providing new insights into the structural plasticity of the matrix during maturation and its functional role in the viral lifecycle.

Guanylate-binding protein (GBP) 5 is an interferon-inducible cellular factor with broad anti-viral activity. Recently, GBP5 has been shown to antagonize the glycoproteins of a number of enveloped viruses, in part by disrupting the host enzyme furin. Here we show that GBP5 strongly impairs the infectivity of virus particles bearing not only viral glycoproteins that depend on furin cleavage for infectivity-the envelope (Env) glycoproteins of HIV-1 and murine leukemia virus and the spike (S) glycoprotein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-but also viral glycoproteins that do not depend on furin cleavage: vesicular stomatitis virus glycoprotein and SARS-CoV S. We observe that GBP5 disrupts proper N -linked protein glycosylation and reduces the incorporation of viral glycoproteins into virus particles. The glycosylation of the cellular protein CD4 is also altered by GBP5 expression. Flow cytometry analysis shows that GBP5 expression reduces the cell-surface levels of HIV-1 Env and the S glycoproteins of SARS-CoV and SARS-CoV-2. Our data demonstrate that, under the experimental conditions used, inhibition of furin-mediated glycoprotein cleavage is not the primary anti-viral mechanism of action of GBP5. Rather, the antagonism appears to be related to impaired trafficking of glycoproteins to the plasma membrane. These results provide novel insights into the broad antagonism of viral glycoprotein function by the cellular host innate immune response., Importance: The surface of enveloped viruses contains viral envelope glycoproteins, an important structural component facilitating virus attachment and entry while also acting as targets for the host adaptive immune system. In this study, we show that expression of GBP5 in virus-producer cells alters the glycosylation, cell-surface expression, and virion incorporation of viral glycoproteins across several virus families. This research provides novel insights into the broad impact of the host cell anti-viral factor GBP5 on protein glycosylation and trafficking., Competing Interests: The authors declare no conflict of interest.

Sleep and inflammatory cytokines have a bidirectional relationship where circadian rhythms influence increase in levels of certain cytokines, and in return, some cytokines induce sleep, as we frequently experience during illnesses. The most commonly studied cytokines, in the context of inflammation and sleep, are interleukin 6 (IL-6), tumor necrosis factor (TNF), and (IL-1). In this article, the author follows the effect of circadian rhythms on blood levels of these cytokines and explores the changes in their levels in conditions that affect sleep, such as obstructive sleep apnea and insomnia., (Copyright © 2023. Published by Elsevier Inc.)

Objective: To characterize the clinical characteristics of infants with obstructive sleep apnea (OSA), define the resolution rate of infant OSA, and identify factors associated with OSA resolution., Methods: We identified infants diagnosed with OSA via retrospective chart review at less than one year of age at a tertiary care center. We identified patient comorbidities, flexible or rigid airway evaluations, surgical procedures, and oxygen/other respiratory support administration. We identified infants as having resolved OSA based on clinical or polysomnogram resolution. We compared the frequency of comorbid diagnoses and receipt of interventions in infants with resolved versus non-resolved OSA by χ 2 analysis., Results: 83 patients were included. Prematurity was found in 35/83 (42%), hypotonia-related diagnoses in 31/83 (37%), and craniofacial abnormalities in 34/83 (41%). Resolution was observed in 61/83 (74%), either clinically or by polysomnogram, during follow up. On χ 2 analysis, surgical intervention was not associated with likelihood of resolution (73% versus 74% in those without surgical intervention, p = 0.98). Patients with airway abnormalities on flexible or rigid evaluation were less likely to have OSA resolution than those without (63% versus 100%, p = 0.010), as were patients with hypotonia-related diagnoses (58% versus 83%, p = 0.014). In patients with laryngomalacia, there was no association of supraglottoplasty with increased resolution (88% with supraglottoplasty versus 80% without, p = 1.00)., Conclusions: We identified a group of infants with OSA with diverse comorbidities. There was a high rate of resolution. This data can assist with treatment planning and family counselling for infants with OSA. A prospective clinical trial is needed to better assess consequences of OSA in this age., Competing Interests: Declaration of competing interest None., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Study Objectives: Craniofacial malformations with micrognathia cause high grades of obstructive sleep apnea (OSA) measured by polysomnography (PSG). Mandibular distraction osteogenesis is a novel procedure for upper airway obstruction relief. Our primary objective was to describe the utilization of PSGs to improve obstruction in patients undergoing mandibular distraction., Methods: This is a retrospective study. Patients with micrognathia and severe upper airway obstruction, presenting with severe OSA diagnosed by PSG, were included from a single tertiary care center between 2015 and 2019. PSGs were done (1) prior to surgery, (2) once the cosmetic goal was achieved (Post-Op 1), and (3) if residual moderate-to-severe OSA was seen, every 2 nights until mild or no OSA was achieved (Post-Op 2)., Results: Thirteen patients were included. The median age at surgery was 1.1 months (10 days-3 months). All 13 patients had baseline severe OSA, with a median obstructive apnea-hypopnea index of 33 events/h and a median O 2 nadir of 73%. Post-Op 1 PSG was done at a median of 6 days after surgery. Median first postoperative obstructive apnea-hypopnea index in all 13 patients was 6.8 events/h, with a median O 2 nadir of 87%. A median additional distraction of 3 mm was needed beyond the traditionally recommended advancement. Long-term follow-up studies at or after 1 year were done in 5 patients, all showing persistent nonsevere OSA., Conclusions: This is the first case series utilizing PSGs as a guide for mandibular distraction osteogenesis in patients with micrognathia showing the need for jaw overcorrection to achieve resolution of OSA., Citation: Kochhar R, Modi V, de Silva N, et al. Polysomnography-guided mandibular distraction osteogenesis in Pierre Robin sequence patients. J Clin Sleep Med . 2022;18(7):1749-1755., (© 2022 American Academy of Sleep Medicine.)

Influenza A virus (IAV) contains a segmented RNA genome that is transcribed and replicated by the viral RNA polymerase in the cell nucleus. Replicated RNA segments are assembled with viral polymerase and oligomeric nucleoprotein into viral ribonucleoprotein (vRNP) complexes which are exported from the nucleus and transported across the cytoplasm to be packaged into progeny virions. Host GTPase Rab11a associated with recycling endosomes is believed to contribute to this process by mediating the cytoplasmic transport of vRNPs. However, how vRNPs interact with Rab11a remains poorly understood. In this study, we utilized a combination of biochemical, proteomic, and biophysical approaches to characterize the interaction between the viral polymerase and Rab11a. Using pulldown assays, we showed that vRNPs but not complementary RNPs (cRNPs) from infected cell lysates bind to Rab11a. We also showed that the viral polymerase directly interacts with Rab11a and that the C-terminal two-thirds of the PB2 polymerase subunit (PB2-C) comprising the cap-binding, mid-link, 627, and nuclear localization signal (NLS) domains mediate this interaction. Small-angle X-ray scattering (SAXS) experiments confirmed that PB2-C associates with Rab11a in solution forming a compact folded complex with a 1:1 stoichiometry. Furthermore, we demonstrate that the switch I region of Rab11a, which has been shown to be important for binding Rab11 family-interacting proteins (Rab11-FIPs), is also important for PB2-C binding, suggesting that IAV polymerase and Rab11-FIPs compete for the same binding site. Our findings expand our understanding of the interaction between the IAV polymerase and Rab11a in the cytoplasmic transport of vRNPs. IMPORTANCE The influenza virus RNA genome segments are replicated in the cell nucleus and are assembled into viral ribonucleoprotein (vRNP) complexes with viral RNA polymerase and nucleoprotein (NP). Replicated vRNPs need to be exported from the nucleus and trafficked across the cytoplasm to the cell membrane, where virion assembly takes place. The host GTPase Rab11a plays a role in vRNP trafficking. In this study, we showed that the viral polymerase directly interacts with Rab11a mediating the interaction between vRNPs and Rab11a. We mapped this interaction to the C-terminal domains of the PB2 polymerase subunit and the switch I region of Rab11a. Identifying the exact site of Rab11a binding on the viral polymerase could uncover a novel target site for the development of an influenza antiviral drug.

Study Objectives: (1) To determine the characteristics of term and preterm infants for whom polysomnography (PSG) was used as a primary diagnostic tool in infants with recurrent desaturation episodes, suspected obstructive apnea, or both, and the prevalence of abnormal studies. (2) To identify the interventions following PSGs. (3) To assess the added value of airway and swallow evaluations., Methods: Retrospective cohort study of infants evaluated by PSG in the Neonatal Intensive Care Unit at New York-Presbyterian Hospital-Weill Cornell from January 2012 to April 2018., Results: PSGs were performed on 31 infants; 15 (48%) term and 16 (52%) preterm infants. Indications for PSG were persistent desaturations (n = 24), suspected obstructive apnea (n = 15), and stridor (n = 2). Primary comorbid conditions were respiratory (n = 11), craniofacial (n = 9), airway anomalies (n = 6), and neurologic (n = 5). The apnea-hypopnea index was abnormal in 30 (97%) infants. Of those, 23 (74%) were severe, 7 (23%) were moderate, and 1 was normal (3%). Apneic events were predominantly obstructive in 23 infants and predominantly central in 6. The apnea-hypopnea index improved in all but 1 follow-up PSG. The PSG findings resulted in interventions in 24 (77%) infants, in addition to concomitant otolaryngology evaluations (abnormal in 20/25) and swallow studies (abnormal in 9/14). Clinical signs completely resolved in 22 (71%) infants., Conclusions: This is one of the first reports on the diagnostic value of inpatient PSGs in the neonatal intensive care unit in infants with recurrent desaturation episodes, suspected obstructive apnea, or both. Our findings indicate that PSG is an important tool in evaluating and targeting therapies in complex term and preterm infants with a wide variety of comorbidities., Citation: Kim J, Gueye-Ndiaye S, Mauer E, Modi VK, Perlman J, Veler H. Polysomnography use in complex term and preterm infants to facilitate evaluation and management in the neonatal intensive care unit. J Clin Sleep Med . 2021;17(8):1653-1663., (© 2021 American Academy of Sleep Medicine.)

Fungi can exacerbate symptoms in patients with asthma. To our knowledge, genetic risk factors for fungal-associated asthma have not been described. We present here the cases of 6 children who carried the diagnosis of severe asthma with fungal sensitization, 3 of whom were treated with and responded clinically to itraconazole therapy. All 6 patients were heterozygous for a 24-base pair duplication in the CHIT1 gene, which has been associated with decreased levels of circulating chitotriosidase and susceptibility to fungal infection.

Although rigid bronchoscopy is the procedure of choice for interventional procedures of the proximal airway, flexible bronchoscopy can be used when lesions are not accessible by rigid equipment. We present an adolescent patient with tracheal stenosis whose airway was inaccessible through rigid bronchoscopy and thus required flexible bronchoscopy for all therapeutic procedures, including a stent placement. In addition, we describe our use of impulse oscillometry to monitor stent patency.

Objective: Demonstrate the benefits of a multidisciplinary pediatric airway team prepared to evaluate and treat otolaryngology patients with flexible bronchoscopy., Design: Case series., Setting: Tertiary, academic children's hospital., Patients: 10 children (5 male, 5 female age range 2 months-16 years) presenting with complex symptoms potentially referable to large airways., Intervention: Flexible bronchoscopy for diagnostic (bronchoalveolar lavage, ciliary biopsy, assess ongoing surgical intervention, and rule in or rule out foreign body; N=6) or therapeutic (evacuate bronchial mucus plug, laser subglottis when patient has fused cervical spine, and distal instillation [fibrin glue for bronchopleural fistula and dornase alpha for plastic bronchitis]; N=4)., Main Outcome Measure: Retrospectively ask if flexible bronchoscopy and interdisciplinary management improved patient care in these select otolaryngology cases., Results: 10/10 patients benefited from interdisciplinary management including flexible bronchoscopy., Conclusion: Our experience illustrates many uses for flexible bronchoscopy in otolaryngology patients, and suggests that an airway team prepared to use flexible bronchoscopy will create opportunities for improved patient care.

Microbial products serving as superantigens (SAgs) have been implicated in triggering various T cell-mediated chronic inflammatory disorders, including severe asthma. Given earlier evidence demonstrating that airway smooth muscle (ASM) cells express MHC class II molecules, we investigated whether ASM can present SAg to resting CD4(+) T cells, and further examined whether this action reciprocally elicits proasthmatic changes in ASM responsiveness. Coincubation of CD4(+) T cells with human ASM cells pulsed with the SAg, staphylococcal enterotoxin A (SEA), elicited adherence and clustering of class II and CD3 molecules at the ASM/T cell interface, indicative of immunological synapse formation, in association with T cell activation. This ASM/T cell interaction evoked up-regulated mRNA expression and pronounced release of the Th2-type cytokine, IL-13, into the coculture medium, which was MHC class II dependent. Moreover, when administering the conditioned medium from the SEA-stimulated ASM/T cell cocultures to isolated naive rabbit ASM tissues, the latter exhibited proasthmatic-like changes in their constrictor and relaxation responsiveness that were prevented by pretreating the tissues with an anti-IL-13 neutralizing Ab. Collectively, these observations are the first to demonstrate that ASM can present SAg to CD4(+) T cells, and that this MHC class II-mediated cooperative ASM/T cell interaction elicits release of IL-13 that, in turn, evokes proasthmatic changes in ASM constrictor and relaxant responsiveness. Thus, a new immuno-regulatory role for ASM is identified that potentially contributes to the pathogenesis of nonallergic (intrinsic) asthma and, accordingly, may underlie the reported association between microbial SAg exposure, T cell activation, and severe asthma.

Activation of Toll-like receptors (TLRs) on immune surveillance cells in the lung has been implicated in the pathobiology of allergic asthma, a condition associated with altered airway smooth muscle (ASM) contractility. Because ASM is known to directly respond to various proasthmatic stimuli, the potential role of TLR signaling in ASM in regulating airway expression of the proasthmatic phenotype was investigated. Cultured human ASM cells were found to express TLR4 and TLR9 mRNA transcripts and, whereas TLR9 stimulation had little effect, TLR4 activation with LPS elicited significant increases in IL-6 release and evoked proasthmatic-like changes in the constrictor and relaxation responsiveness of isolated rabbit ASM tissues. Complementary studies further demonstrated that the ASM responses to LPS were associated with activation of the ERK1/2 and p38 MAPK signaling pathways, IKK-mediated activation of NF-kappaB, and coupling of phosphorylated ERK1/2 with the p65 subunit of NF-kappaB. Moreover, the induced NF-kappaB activity and changes in ASM responsiveness were prevented in LPS-exposed ASM that were pretreated with inhibitors of ERK1/2 signaling, whereas inhibition of p38 MAPK augmented the proasthmatic responses to LPS. Finally, activation of p38 MAPK with anisomycin prevented both the LPS-induced stimulation of ERK1/2-mediated NF-kappaB activity and associated changes in ASM responsiveness. Collectively, these data support the novel concept that TLR4 activation in ASM elicits changes in ASM function that are regulated by opposing effects of MAPK signaling, wherein LPS-induced ERK1/2 activation mediates NF-kappaB-dependent proasthmatic-like changes in ASM function, whereas coactivation of p38 MAPK serves to homeostatically downregulate the proasthmatic effects of ERK1/2 activation.

Background: House dust mite allergen exposure is a key risk factor for the development of allergic asthma. Beyond provoking immune cell-mediated allergic responses, house dust mite allergens were recently shown to exert direct effects on airway structural cells secondary to their intrinsic protease activities., Objective: This study tested the hypothesis that house dust mite allergen exposure can produce changes in airway responsiveness through a direct effect on airway smooth muscle (ASM)., Methods: Isolated rabbit ASM tissues were exposed to the house dust mite allergen, Der p 1, and induced changes in ASM responsiveness and activation of mitogen-activated protein kinase (MAPK) signaling pathways were examined under different experimental conditions., Results: The observations demonstrated the following: (1) Der p 1 exposure elicited enhanced constrictor responses and impaired relaxation responses in the ASM tissues, (2) these proasthmatic-like effects of Der p 1 were attributed to its intrinsic cysteine protease activity, and (3) the induced changes in ASM responsiveness were associated with activation of both the extracellular signal-regulated kinase (ERK) 1/2 and the p38 MAPK signaling pathways. Additionally, specific blockade of ERK1/2 signaling was found to prevent the Der p 1-induced changes in ASM responsiveness, whereas inhibition of p38 MAPK signaling enhanced the proasthmatic-like action of Der p 1, with the latter effect a result of augmented activation of ERK1/2., Conclusion: These findings are the first to demonstrate that the dust mite allergen, Der p 1, can directly elicit changes in ASM responsiveness that are associated with activation of MAPK signaling, wherein proasthmatic effects induced by Der p 1 are attributed to activation of ERK1/2, whereas coactivation of p38 MAPK exerts a homeostatic action by negatively regulating ERK1/2 signaling.