The Association of Infant Urinary Adrenal Steroids with the Risk of Childhood Asthma Development.

BACKGROUND: Adrenal steroids play important roles in early life development. However, our understanding of the effects of perinatal adrenal steroids on the development of childhood asthma is incomplete. OBJECTIVE: This study aimed to evaluate the associations between early life adrenal steroid levels and childhood asthma. METHODS: Participants included INSPIRE birth cohort children with untargeted urinary metabolomics data measured during early infancy (n=264) and nasal immune mediator data measured concurrently at age 2-6 months (n=76). A total of 11 adrenal steroid compounds identified using untargeted metabolomics and 6 asthma-relevant nasal immune mediators from multiplex assays were a priori selected. Current asthma at ages 5 and 6 years was ascertained using validated questionnaires. Associations were tested using logistic and linear regression with confounders adjustment. RESULTS: Pregnenetriol disulfate (adjusted odd ratio, aOR= 0.20, 95%CI= [0.06, 0.68]) and 3a,21-dihydroxy-5b-pregnane-11,20-dione-21-glucuronide (aOR= 0.34, 95%CI= [0.14, 0.75]) were inversely associated with childhood asthma at 5 and 6 years after multiple testing adjustment. There was a significant interaction effect of pregnanediol-3-glucuronide by sex (aOR=0.11, 95%CI=[0.02, 0.51], for females) on childhood asthma. Pregnenetriol disulfate was inversely associated with GM-CSF (b=-0.45, q-value=0.05). 3a,21-dihydroxy-5b-pregnane-11,20-dione 21-glucuronide was inversely associated with IL-4 (b=-0.29, q-value=0.02), IL-5 (b=-0.35, q-value=0.006), IL-13 (b=-0.26, q-value=0.02), GM-CSF (b=-0.35, q-value=0.006), and FGF-b(b=-0.24, q-value=0.01) after multiple testing adjustment. CONCLUSION: The inverse association between adrenal steroids downstream of progesterone and 17-hydroxypregnenolone and odds of childhood asthma and nasopharyngeal Type 2 immune biomarkers suggest that increased early life adrenal steroids may suppress Type 2 inflammation and protect against the development of childhood asthma.

Exploring the role of CITED transcriptional regulators in the control of macrophage polarization.

Macrophages are tissue resident innate phagocytic cells that take on contrasting phenotypes, or polarization states, in response to the changing combination of microbial and cytokine signals at sites of infection. During the opening stages of an infection, macrophages adopt the proinflammatory, highly antimicrobial M1 state, later shifting to an anti-inflammatory, pro-tissue repair M2 state as the infection resolves. The changes in gene expression underlying these transitions are primarily governed by nuclear factor kappaB (NF-κB), Janus kinase (JAK)/signal transducer and activation of transcription (STAT), and hypoxia-inducible factor 1 (HIF1) transcription factors, the activity of which must be carefully controlled to ensure an effective yet spatially and temporally restricted inflammatory response. While much of this control is provided by pathway-specific feedback loops, recent work has shown that the transcriptional co-regulators of the CBP/p300-interacting transactivator with glutamic acid/aspartic acid-rich carboxy-terminal domain (CITED) family serve as common controllers for these pathways. In this review, we describe how CITED proteins regulate polarization-associated gene expression changes by controlling the ability of transcription factors to form chromatin complexes with the histone acetyltransferase, CBP/p300. We will also cover how differences in the interactions between CITED1 and 2 with CBP/p300 drive their contrasting effects on pro-inflammatory gene expression.

Effects of end-stage osteoarthritis on markers of skeletal muscle Long INterspersed Element-1 activity.

OBJECTIVE: Long INterspersed Element-1 (L1) is an autonomous transposable element in the genome. L1 transcripts that are not reverse transcribed back into the genome can accumulate in the cytoplasm and activate an inflammatory response via the cyclic GMP-AMP (cGAS)-STING pathway. We examined skeletal muscle L1 markers as well as STING protein levels in 10 older individuals (63 ± 11 y, BMI = 30.2 ± 6.8 kg/m2) with end-stage osteoarthritis (OA) undergoing total hip (THA, n = 4) or knee (TKA, n = 6) arthroplasty versus 10 young, healthy comparators (Y, 22 ± 2 y, BMI = 23.2 ± 2.5 kg/m2). For OA, muscle was collected from surgical (SX) and contralateral (CTL) sides whereas single vastus lateralis samples were collected from Y. RESULTS: L1 mRNA was higher in CTL and SX compared to Y (p < 0.001 and p = 0.001, respectively). Protein expression was higher in SX versus Y for ORF1p (p = 0.002) and STING (p = 0.022). While these data are preliminary due to limited n-sizes and the lack of a BMI-matched younger control group, higher L1 mRNA expression, ORF1p and STING protein are evident in older versus younger adults. More research is needed to determine whether cGAS-STING signaling contributes to heightened muscle inflammation during aging and/or OA.

Exclusive breast-feeding, the early-life microbiome and immune response, and common childhood respiratory illnesses.

BACKGROUND: The impact of breast-feeding on certain childhood respiratory illnesses remains controversial. OBJECTIVE: We sought to examine the effect of exclusive breast-feeding on the early-life upper respiratory tract (URT) and gut microbiome, the URT immune response in infancy, and the risk of common pediatric respiratory diseases. METHODS: We analyzed data from a birth cohort of healthy infants with prospective ascertainment of breast-feeding patterns and common pediatric pulmonary and atopic outcomes. In a subset of infants, we also characterized the URT and gut microbiome using 16S ribosomal RNA sequencing and measured 9 URT cytokines using magnetic bead-based assays. RESULTS: Of the 1949 infants enrolled, 1495 (76.71%) had 4-year data. In adjusted analyses, exclusive breast-feeding (1) had an inverse dose-response on the ⍺-diversity of the early-life URT and gut microbiome, (2) was positively associated with the URT levels of IFN-α, IFN-γ, and IL-17A in infancy, and (3) had a protective dose-response on the development of a lower respiratory tract infection in infancy, 4-year current asthma, and 4-year ever allergic rhinitis (odds ratio [95% CI] for each 4 weeks of exclusive breast-feeding, 0.95 [0.91-0.99], 0.95 [0.90-0.99], and 0.95 [0.92-0.99], respectively). In exploratory analyses, we also found that the protective association of exclusive breast-feeding on 4-year current asthma was mediated through its impact on the gut microbiome (P = .03). CONCLUSIONS: Our results support a protective causal role of exclusive breast-feeding in the risk of developing a lower respiratory tract infection in infancy and asthma and allergic rhinitis in childhood. They also shed light on potential mechanisms of these associations, including the effect of exclusive breast-feeding on the gut microbiome.

Perioperative assessment of muscle inflammation susceptibility in patients with end-stage osteoarthritis.

Many individuals with end-stage osteoarthritis (OA) undergo elective total hip/knee arthroplasty (THA/TKA) to relieve pain, improve mobility and quality of life. However, ∼30% suffer long-term mobility impairment following surgery. This may be in part due to muscle inflammation susceptibility (MuIS+), an overt proinflammatory pathology localized to skeletal muscle surrounding the diseased joint, present in some patients with TKA/THA. We interrogated the hypothesis that MuIS+ status results in a perturbed perioperative gene expression profile and decreases skeletal muscle integrity in patients with end-stage OA. Samples were leveraged from the two-site, randomized, controlled trial R01HD084124, NCT02628795. Participants were dichotomized based on surgical (SX) muscle gene expression of TNFRSF1A (TNF-αR). MuIS+/- samples were probed for gene expression and fibrosis. Paired and independent two-tailed t tests were used to determine differences between contralateral (CTRL) and surgical (SX) limbs and between-subject comparisons, respectively. Significance was declared at P < 0.05. Seventy participants (26M/44F; mean age 62.41 ± 8.86 yr; mean body mass index 31.10 ± 4.91 kg/m2) undergoing THA/TKA were clustered as MuIS+ (n = 24) or MuIS- (n = 46). Lower skeletal muscle integrity (greater fibrosis) exists on the SX versus CTRL limb (P < 0.001). Furthermore, MuIS+ versus MuIS- muscle exhibited higher proinflammatory (IL-6R and TNF-α) and catabolic (TRIM63) gene expression (P < 0.001, P = 0.004, and 0.024 respectively), with a trend for greater fibrosis (P = 0.087). Patients with MuIS+ exhibit more inflammation and catabolic gene expression in skeletal muscle of the SX limb, accompanied by decreased skeletal muscle integrity (Trend). This highlights the impact of MuIS+ status emphasizing the potential value of perioperative MuIS assessment to inform optimal postsurgical care.NEW & NOTEWORTHY This study assessed the skeletal muscle molecular characteristics associated with end-stage osteoarthritis and refined an important phenotype, in some patients, termed muscle inflammation susceptibility (MuIS+) that may be an important consideration following surgery. Furthermore, we provide evidence of differential inflammatory and catabolic gene expression between the contralateral and surgical limbs along with differences between the skeletal muscle surrounding the diseased hip versus knee joints.

Cellular and systemic energy metabolic dysregulation in asthma development-a hypothesis-generating approach.

BACKGROUND: The roles of systemic and airway-specific epithelial energy metabolism in altering the developmental programming of airway epithelial cells (AECs) in early life are poorly understood. OBJECTIVE: Our aim was to assess carbohydrate metabolism in developing AECs among children with and without wheeze and test the association of infant plasma energy biomarkers with subsequent recurrent wheeze and asthma outcomes. METHODS: We measured cellular carbohydrate metabolism in live nasal AECs collected at age 2 years from 15 male subjects with and without a history of wheeze and performed a principal component analysis to visually assess clustering of data on AEC metabolism of glycolitic metabolites and simple sugars. Among 237 children with available year 1 plasma samples, we tested the associations of year 1 plasma energy biomarkers and recurrent wheeze and asthma by using generalized estimating equations and logistic regression. RESULTS: Children with a history of wheeze had lower utilization of glucose in their nasal AECs than did children with no wheeze. Systemically, a higher plasma glucose concentration at year 1 (within the normal range) was associated with decreased odds of asthma at age 5 years (adjusted odds ratio = 0.56; 95% CI = 0.35-0.90). Insulin concentration, glucose-to-insulin ratio, C-peptide concentration, and leptin concentration at year 1 were associated with recurrent wheeze from age 2 years to age 5 years. CONCLUSION: These results suggest that there is significant energy metabolism dysregulation in early life, which likely affects AEC development. These pertubations of epithelial cell metabolism in infancy may have lasting effects on lung development that could render the airway more susceptible to allergic sensitization.

Upper respiratory tract bacterial-immune interactions during respiratory syncytial virus infection in infancy.

BACKGROUND: The risk factors determining short- and long-term morbidity following acute respiratory infection (ARI) due to respiratory syncytial virus (RSV) in infancy remain poorly understood. OBJECTIVES: Our aim was to examine the associations of the upper respiratory tract (URT) microbiome during RSV ARI in infancy with the acute local immune response and short- and long-term clinical outcomes. METHODS: We characterized the URT microbiome by 16S ribosomal RNA sequencing and assessed the acute local immune response by measuring 53 immune mediators with high-throughput immunoassays in 357 RSV-infected infants. Our short- and long-term clinical outcomes included several markers of disease severity and the number of wheezing episodes in the fourth year of life, respectively. RESULTS: We found several specific URT bacterial-immune mediator associations. In addition, the Shannon ⍺-diversity index of the URT microbiome was associated with a higher respiratory severity score (ß =.50 [95% CI = 0.13-0.86]), greater odds of a lower ARI (odds ratio = 1.63 [95% CI = 1.10-2.43]), and higher number of wheezing episodes in the fourth year of life (ß = 0.89 [95% CI = 0.37-1.40]). The Jaccard ß-diversity index of the URT microbiome differed by level of care required (P = .04). Furthermore, we found an interaction between the Shannon ⍺-diversity index of the URT microbiome and the first principal component of the acute local immune response on the respiratory severity score (P = .048). CONCLUSIONS: The URT microbiome during RSV ARI in infancy is associated with the acute local immune response, disease severity, and number of wheezing episodes in the fourth year of life. Our results also suggest complex URT bacterial-immune interactions that can affect the severity of the RSV ARI.

Evaluation of the upper airway microbiome and immune response with nasal epithelial lining fluid absorption and nasal washes.

Despite being commonly used to collect upper airway epithelial lining fluid, nasal washes are poorly reproducible, not suitable for serial sampling, and limited by a dilution effect. In contrast, nasal filters lack these limitations and are an attractive alternative. To examine whether nasal filters are superior to nasal washes as a sampling method for the characterization of the upper airway microbiome and immune response, we collected paired nasal filters and washes from a group of 40 healthy children and adults. To characterize the upper airway microbiome, we used 16S ribosomal RNA and shotgun metagenomic sequencing. To characterize the immune response, we measured total protein using a BCA assay and 53 immune mediators using multiplex magnetic bead-based assays. We conducted statistical analyses to compare common microbial ecology indices and immune-mediator median fluorescence intensities (MFIs) between sample types. In general, nasal filters were more likely to pass quality control in both children and adults. There were no significant differences in microbiome community richness, α-diversity, or structure between pediatric samples types; however, these were all highly dissimilar between adult sample types. In addition, there were significant differences in the abundance of amplicon sequence variants between sample types in children and adults. In adults, total proteins were significantly higher in nasal filters than nasal washes; consequently, the immune-mediator MFIs were not well detected in nasal washes. Based on better quality control sequencing metrics and higher immunoassay sensitivity, our results suggest that nasal filters are a superior sampling method to characterize the upper airway microbiome and immune response in both children and adults.

The vitamin D activator CYP27B1 is upregulated in muscle fibers in denervating disease and can track progression in amyotrophic lateral sclerosis.

Extra-renal expression of Cytochrome P450 Family 27 Subfamily B Member 1 (CYP27B1) has been well recognized and reflects the importance of intracrine/paracrine vitamin D signaling in different tissues under physiological and pathological conditions. In a prior RNA sequencing project, we identified CYP27B1 mRNA as upregulated in muscle samples from patients with amyotrophic lateral sclerosis (ALS) compared to normal controls. Our aims here were: (1) to validate this finding in a larger sample set including disease controls, (2) to determine which cell type is expressing CYP27B1 protein in muscle tissue, (3) to correlate CYP27B1 mRNA expression with disease progression in the SOD1G93A ALS mouse and in ALS patients. We assessed CYP27B1 expression by qPCR, western blot, and immunohistochemistry in a repository of muscle samples from ALS, disease controls (myopathy and non-ALS neuropathic disease), normal subjects, and muscle samples from the SOD1G93A mouse. Eight ALS patients were studied prospectively over 6-12 months with serial muscle biopsies. We found that CYP27B1 mRNA and protein levels were significantly increased in ALS versus normal and myopathy muscle samples. Neuropathy samples had increased CYP27B1 mRNA and protein expression but at a lower level than the ALS group. Immunohistochemistry showed that CYP27B1 localized to myofibers, especially those with features of denervation. In the SOD1G93A mouse, CYP27B1 mRNA and protein were detected in skeletal muscle in early pre-symptomatic stages and increased through end-stage. In the human study, increases in CYP27B1 mRNA in muscle biopsies correlated with disease progression rates over the same time period. In summary, we show for the first time that CYP27B1 mRNA and protein expression are elevated in muscle fibers in denervating disease, especially ALS, where mRNA levels can potentially serve as a surrogate marker for tracking disease progression. Its upregulation may reflect a local perturbation of vitamin D signaling, and further characterization of this pathway may provide insight into underlying molecular processes linked to muscle denervation.