Effects of highly effective modulator therapy on the dynamics of the respiratory mucosal environment and inflammatory response in cystic fibrosis.

BACKGROUND: While the widespread initiation of elexacaftor/tezacaftor/ivacaftor (ETI) has led to dramatic clinical improvements among persons with cystic fibrosis (pwCF), little is known about how ETI affects the respiratory mucosal inflammatory and physiochemical environment, or how these changes relate to lung function. METHODS: We performed a prospective, longitudinal study of adults with CF and chronic rhinosinusitis (CF-CRS) followed at our CF center (n = 18). Endoscopic upper respiratory tract (paranasal sinus) aspirates from multiple visit dates, both pre- and post-ETI initiation, were collected and tested for cytokines, metals, pH, and lactate levels. Generalized estimating equations were used to identify relationships between ETI and upper respiratory tract (URT) biomarker levels, and between URT biomarkers and lung function or clinical sinus parameters. RESULTS: ETI was associated with decreased upper respiratory mucosal cytokines B-cell activating factor (BAFF), IL-12p40, IL-32, IL-8, IL-22 and soluble tumor necrosis factor-1 (sTNFR1), and an increase in a proliferation-inducing ligand (APRIL) and IL-19. ETI was also associated with decreased URT levels of copper, manganese, and zinc. In turn, lower URT levels of BAFF, IL-8, lactate, and potassium were each associated with ~1.5% to 4.3% improved forced expiratory volume in 1 s (FEV1), while higher levels of IFNγ, iron, and selenium were associated with ~2% to 10% higher FEV1. CONCLUSIONS: Our observations suggest a dampening of inflammatory signals and restriction in microbial nutrients in the upper respiratory tract with ETI. These findings improve our understanding of how ETI impacts the mucosal environment in the respiratory tract, and may give insight into the improved infectious and inflammatory status and the resulting clinical improvements seen in pwCF.

Local cytokine levels associate with SNOT-22 and UPSIT scores in chronic rhinosinusitis.

KEY POINTS: Elevated IL-5, IL-13, IL-33, and CCL2 correlate with lower UPSIT scores in CRS and AERD patients. Elevated IL-5, IL-13, TNF-α, CCL2, and CXCL-8 correlate with higher SNOT-22 scores in CRS and AERD patients.

Olfactory and neurological outcomes of SARS-CoV-2 from acute infection to recovery.

Educational objective: To investigate the impact of SARS-CoV-2 on sinonasal quality of life, olfaction, and cognition at different stages of viral infection and evaluate the association between olfaction and cognition in this population cohort. Objectives: While olfactory dysfunction (OD) is a frequently reported symptom of COVID-19 (98% prevalence), neurocognitive symptoms are becoming more apparent as patients recover from infection. This study aims to address how different stages of infection [active infection (positive PCR test, symptomatic) vs. recovered (7 days post-symptoms)] compared to healthy control patients influence sinonasal quality of life, olfactory function, and cognition. Study design: Prospective, longitudinal, case-control. Methods: Participants completed the SNOT-22, University of Pennsylvania Smell Identification Test (UPSIT) and validated cognitive examinations to assess degree of smell loss and neurocognitive function at baseline and at 1 and 3 months for the active group and 3 months for the recovered group. Self-reported olfactory function and overall health metrics were also collected. Results: The recovered group had the lowest average UPSIT score of 27.6 compared to 32.7 (active) and 32.6 (healthy control). 80% (n = 24) of the recovered patients and 56.3% (n = 9) of the active patients suffered from smell loss. In follow-up, the active group showed improvement in UPSIT scores while the recovered group scores worsened. In terms of neurocognitive performance, recovered patients had lower processing speed despite an improving UPSIT score. Conclusion: SARS-CoV-2 infection was found to impact olfactory function in a delayed fashion with significant impact despite recovery from active infection. Although olfactory function improved, decrements in cognitive processing speed were detected in our cohort.

Fractional Exhaled Nitric Oxide as a Marker of Mucosal Inflammation in Chronic Rhinosinusitis.

BACKGROUND: Fractional exhaled nitric oxide (FeNO) is a cost-effective, noninvasive point-of-care test that has proven valuable in identifying patients with lower airway inflammation and predicting the likelihood of responsiveness to inhaled corticosteroid therapy in asthma. The utility of FeNO in upper airway disease, specifically in CRS, remains to be determined. OBJECTIVE: The goal of this study was to test whether FeNO could serve as a noninvasive marker of sinonasal mucosal inflammation in CRS patients. METHODS: FeNO was obtained using a nitric oxide analyzer (NIOX VERO) as well as nasal mucus, the 22-item Sinonasal Outcome Test (SNOT-22), University of Pennsylvania Smell Identification Test (UPSIT), and Lund-Kennedy endoscopic scores concurrently in 112 CRS patients. Nasal mucus was analyzed for cytokine expression using solid-phase sandwich ELISA. Linear regression with Spearman correlation coefficient was used to determine strength of relationship between variables. RESULTS: CRS patients showed elevated FeNO levels with asthma (47.12 ± 5.21 ppb) or without asthma (43.24 ± 9.810 ppb). Elevated FeNO levels correlated with sinonasal mucosal inflammation, as determined by increased levels of CCL26 and TNFα in nasal mucus obtained from CRS patients. Furthermore, elevated FeNO levels selectively correlated with worsened SNOT-22 nasal symptoms (P = 0.03) and Lund-Kennedy endoscopic scores (P = 0.007), but did not correlate with UPSIT scores. CONCLUSIONS: FeNO levels correlated with increased sinonasal mucosal inflammation and symptom severity in CRS regardless of asthma status. FeNO measurements may serve as a quick and noninvasive marker in evaluating CRS patients.

Relationship between socioeconomic status, exposure to airborne pollutants, and chronic rhinosinusitis disease severity.

BACKGROUND: Air pollution directly interacts with airway mucosa, yet little is known about how pollutants affect upper airway inflammation. Studies have shown increased incidence of chronic rhinosinusitis (CRS), rhinitis, and asthma in areas with higher traffic pollution, and these neighborhoods are often associated with lower socioeconomic status (SES). The Area Deprivation Index (ADI) assesses neighborhood-level SES by zip code. The purpose of this study was to assess the relationship between SES and exposure to inhaled pollutants and CRS disease severity. METHODS: CRS patients with and without nasal polyps (CRSwNP and CRSsNP, respectively) were identified (total patients = 234; CRSwNP patients = 138; CRSsNP patients = 96). Pollutant concentrations, including particulate matter 2.5 (PM2.5 ), black carbon (BC), and nitrogen dioxide (NO2 ), were measured at 70 sites within the defined countywide sites and used to estimate patient exposures. SES was measured by ADI state deciles. Disease severity metrics included the modified Lund-Mackay score (LMS), the need for systemic steroids, and functional endoscopic sinus surgery (FESS). Associations were analyzed and identified using linear, logistic, and Poisson multivariable regression. RESULTS: The distribution of CRSsNP and CRSwNP patients across ADI state deciles was similar. ADI, however, was a predictor of exposure to airborne pollutants (PM2.5 , BC, and NO2 ) with a 1.39%, 2.39%, and 2.49% increase in PM2.5 , BC, and NO2 per increasing decile increment (p < 0.0001), respectively, which demonstrated a direct correlation between deprived neighborhoods and higher levels of exposure to PM2.5 , BC, and NO2 with an increase in pollutant levels per increase in ADI decile. Furthermore, ADI was a predictor for increased steroid treatment. CONCLUSION: Lower SES predicted higher exposure to air pollution and increased disease severity in patients with CRS as demonstrated by the increased need for steroid treatment.

Neuromapping olfactory stimulation using magnetoencephalography - visualizing smell, a proof-of-concept study.

Importance: Currently, clinical assessment of olfaction is largely reliant on subjective methods that require patient participation. The objective method for measuring olfaction, using electroencephalogram (EEG) readings, can be supplemented with the improved temporal resolution of magnetoencephalography (MEG) for olfactory measurement that can delineate cortical and peripheral olfactory loss. MEG provides high temporal and spatial resolution which can enhance our understanding of central olfactory processing compared to using EEG alone. Objective: To determine the feasibility of building an in-house portable olfactory stimulator paired with electrophysiological neuroimaging technique with MEG to assess olfaction in the clinical setting. Design setting and participants: This proof-of-concept study utilized a paired MEG-olfactometer paradigm to assess olfaction in three normosmic participants. We used a two-channel olfactory stimulator to deliver odorants according to a programmed stimulus-rest paradigm. Two synthetic odorants: 2% phenethyl alcohol (rose) and 0.5% amyl acetate (banana) were delivered in increasing increments of time followed by periods of rest. Cortical activity was measured via a 306-channel MEG system. Main outcomes and measures: Primary outcome measure was the relative spectral power for each frequency band, which was contrasted between rest and olfactory stimulation. Results: Compared to rest, olfactory stimulation produced a 40% increase in relative alpha power within the olfactory cortex bilaterally with both odorants. A 25%-30% increase in relative alpha power occurred in the left orbitofrontal cortex and precentral gyrus with phenethyl alcohol stimulation but not amyl acetate. Conclusion and relevance: In this proof-of-concept study, we demonstrate the feasibility of olfactory measurement via an olfactometer-MEG paradigm. We found that odorant-specific cortical signatures can be identified using this paradigm, setting the basis for further investigation of this system as a prognostic tool for olfactory loss.

A primary nasopharyngeal three-dimensional air-liquid interface cell culture model of the pseudostratified epithelium reveals differential donor- and cell type-specific susceptibility to Epstein-Barr virus infection.

Epstein-Barr virus (EBV) is a ubiquitous γ-herpesvirus with latent and lytic cycles. EBV replicates in the stratified epithelium but the nasopharynx is also composed of pseudostratified epithelium with distinct cell types. Latent infection is associated with nasopharyngeal carcinoma (NPC). Here, we show with nasopharyngeal conditionally reprogrammed cells cultured at the air-liquid interface that pseudostratified epithelial cells are susceptible to EBV infection. Donors varied in susceptibility to de novo EBV infection, but susceptible cultures also displayed differences with respect to pathogenesis. The cultures from one donor yielded lytic infection but cells from two other donors were positive for EBV-encoded EBERs and negative for other lytic infection markers. All cultures stained positive for the pseudostratified markers CK7, MUC5AC, α-tubulin in cilia, and the EBV epithelial cell receptor Ephrin receptor A2. To define EBV transcriptional programs by cell type and to elucidate latent/lytic infection-differential changes, we performed single cell RNA-sequencing on one EBV-infected culture that resulted in alignment with many EBV transcripts. EBV transcripts represented a small portion of the total transcriptome (~0.17%). All cell types in the pseudostratified epithelium had detectable EBV transcripts with suprabasal cells showing the highest number of reads aligning to many EBV genes. Several restriction factors (IRF1, MX1, STAT1, C18orf25) known to limit lytic infection were expressed at lower levels in the lytic subcluster. A third of the differentially-expressed genes in NPC tumors compared to an uninfected pseudostratified ALI culture overlapped with the differentially-expressed genes in the latent subcluster. A third of these commonly perturbed genes were specific to EBV infection and changed in the same direction. Collectively, these findings suggest that the pseudostratified epithelium could harbor EBV infection and that the pseudostratified infection model mirrors many of the transcriptional changes imposed by EBV infection in NPC.

Association of air pollutants, airborne occupational exposures, and chronic rhinosinusitis disease severity.

BACKGROUND: Previous work has shown that chronic rhinosinusitis (CRS) severity may be associated with particulate matter 2.5 (PM2.5 ) and black carbon (BC) in CRS patients without nasal polyps (CRSsNP). Data regarding occupational exposures, however, are lacking. We assessed the impact of PM2.5 , BC, as well as occupational airborne exposure on CRS disease severity. METHODS: Patients with CRS with nasal polyps (CRSwNP), CRSsNP, and aspirin-exacerbated respiratory disease (AERD) were identified from an institutionwide database. Spatial modeling from 37 pollutant monitoring sites in Allegheny County was used to estimate exposures. Patient occupations using the 2010 Standard Occupation Classification (SOC10) and airborne occupation exposures to vapors, gases, dusts, fumes, fibers and mists (VGDFFiM) or diesel fumes were recorded. Disease severity was measured by modified Lund-Mackay score (LMS), systemic corticosteroid therapy, and incidence of functional endoscopic sinus surgery (FESS). RESULTS: Two hundred thirty-four patients were included (CRSwNP, n = 113; CRSsNP, n = 96; AERD, n = 25). The prevalence of AERD among those with CRSwNP was 18%. Patients exposed to VGDFFiM or diesel fumes required higher steroid doses vs nonexposed patients (p = 0.015 and p = 0.03, respectively); patients with VGDFFiM levels >5% were more likely to undergo FESS vs nonexposed patients (p = 0.0378). There was no difference in PM2.5 and BC with regard to disease severity and FESS between CRSwNP, CRSsNP, and AERD patients. Steroid use was significantly higher in CRSwNP and AERD vs CRSsNP (p = 0.001). LMS was significantly higher in AERD as compared with CRSwNP and CRSsNP (p = 0.001). CONCLUSION: Occupational airborne exposure to VGDFFiM correlated with increased prevalence of FESS and need for corticosteroids in CRS patients. There was no difference in PM2.5 and BC levels and disease severity outcome measures between CRS subtypes in this subset.

15-Lipoxygenase 1 in nasal polyps promotes CCL26/eotaxin 3 expression through extracellular signal-regulated kinase activation.

BACKGROUND: 15-Lipoxygenase 1 (15LO1) is expressed in airway epithelial cells in patients with type 2-high asthma in association with eosinophilia. Chronic rhinosinusitis with nasal polyps (CRSwNP) is also associated with type 2 inflammation and eosinophilia. CCL26/eotaxin 3 has been reported to be regulated by 15LO1 in lower airway epithelial cells. However, its relation to 15LO1 in patients with CRSwNP or mechanisms for its activation are unclear. OBJECTIVE: We sought to evaluate 15LO1 and CCL26 expression in nasal epithelial cells (NECs) from patients with CRSwNP and healthy control subjects (HCs) and determine whether 15LO1 regulates CCL26 in NECs through extracellular signal-regulated kinase (ERK) activation. METHODS: 15LO1, CCL26, and phosphorylated ERK were evaluated in NECs from patients with CRSwNP and HCs. 15LO1/CCL26 and CCL26/cytokeratin 5 were colocalized by means of immunofluorescence. IL-13-stimulated NECs were cultured at an air-liquid interface with or without 15-lipoxygenase 1 gene (ALOX15) Dicer-substrate short interfering RNAs (DsiRNA) transfection, a specific 15LO1 enzymatic inhibitor, and 2 ERK inhibitors. Expression of 15LO1 and CCL26 mRNA and protein was analyzed by using quantitative RT-PCR, Western blotting, and ELISA. RESULTS: 15LO1 expression was increased in nasal polyp (NP) epithelial cells compared with middle turbinate epithelial cells from patients with CRSwNP and HCs. 15LO1 expression correlated with CCL26 expression and colocalized with CCL26 expression in basal cells of the middle turbinate and NPs from patients with CRSwNP. In primary NECs in vitro, IL-13 induced 15LO1 and CCL26 expression. 15LO1 knockdown and inhibition decreased IL-13-induced ERK phosphorylation and CCL26 expression. ERK inhibition (alone) similarly decreased IL-13-induced CCL26. Phosphorylated ERK expression was increased in NECs from CRSwNP subjects and positively correlated with both 15LO1 and CCL26 expression. CONCLUSIONS: 15LO1 expression is increased in NP epithelial cells and contributes to CCL26 expression through ERK activation. 15LO1 could be considered a novel therapeutic target for CRSwNP.

Impact of high- versus low-risk genotype on sinonasal radiographic disease in cystic fibrosis.

OBJECTIVE: Understanding of how specific mutations impact the cystic fibrosis transmembrane conductance regulator (CFTR) protein has given rise to the classification of CF patients into low-risk and high-risk genotypes. Few prior studies have investigated differences in sinonasal disease between low-risk and high-risk CF genotypes. This multi-institutional review aimed to evaluate radiographic sinus disease severity based on genotype. METHODS: A review was conducted on adult patients with CF evaluated between 2005 to 2017 at three academic institutions. Data including age, gender, CFTR mutation, and presence of a maxillofacial/sinus computed tomography scan was collected. A modified Lund-Mackay score (MLMS) was assigned to each scan, and the presence of sinus aplasia or hypoplasia was determined. Patients were further grouped depending on genotype into low- or high-risk for comparison. RESULTS: A total of 126 patients were included with 99 patients in the high-risk and 21 in the low-risk groups. The high-risk group had significantly higher MLMS than the low-risk group (mean 13.88 vs. 8.06, P < 0.0001, 95% CI -8.196 to -3.462) The rate of frontal (P < 0.01), maxillary (P = 0.04), and sphenoid (P < 0.001) hypoplasia/aplasia was significantly higher in high-risk patients compared to low-risk. CONCLUSION: This is one of the largest studies to date evaluating the impact of CF genotype on paranasal sinus development and disease. Genotype appears to impact sinonasal disease severity and also potentially paranasal sinus cavity development to a degree, although the exact mechanism is unknown. LEVEL OF EVIDENCE: 4 Laryngoscope, 129:788-793, 2019.

TIM-3-Expressing Mast Cells Are Present in Chronic Rhinosinusitis with Nasal Polyps.

Objectives To identify whether TIM-3 expression is present in the mast cell population within nasal polyps and to determine its correlation with clinical severity in patients with chronic rhinosinusitis with nasal polyposis. Study Design Basic science, translational study. Setting Nasal polyp tissue collected from patients seen at a tertiary care hospital (2015-2016). Subjects and Methods Nasal polyp tissue obtained during functional endoscopic sinus surgery (n = 24) was enzymatically digested into epithelial and stromal fractions. Viable mast cells expressing TIM-3 were identified using flow cytometry for the following: CD45, Live/Dead, c-kit, FcεR1, TIM-3. Disease severity was assessed using the Sino-Nasal Outcome Test, Lund-Mackay staging system, Lund-Kennedy staging system, and complete blood counts. Results Mast cells were found in both the epithelial and stromal layers of polyps, with a greater %TIM-3+ mast cells in the epithelial layer compared with that of the stromal layer ( P = .001). As the percentage of mast cells increased, there was a comparative worsening in endoscopic severity after comparing pre- and postoperative LK scores (ρ = -0.455, P = .029). In a subgroup of patients with concomitant asthma, increased epithelial %TIM-3+ mast cells also correlated with worsening endoscopic appearance postoperatively (ρ = 0.866, P = .001, n = 11). Oral corticosteroid treatment did not change the viability of mast cells nor their influence on the increased postoperative endoscopic disease severity (ρ = -0.544, P = .020, n = 18). Conclusion Viable mast cells were found to be present in polyps with increased TIM-3 expression at the epithelial layer. This suggests that TIM-3 may play a role in chronic inflammation in CRSwNP via mast cell activation.

The association of air pollutants and allergic and nonallergic rhinitis in chronic rhinosinusitis.

BACKGROUND: There has been little investigation regarding air quality and rhinitis in the pathophysiology of upper airway disease. In this study, we assessed the impact of inhalant pollutants (particulate matter 2.5 [PM2.5 ] and black carbon [BC]) on allergic rhinitis and chronic rhinosinusitis (CRS) disease severity. METHODS: CRS patients with nasal polyps (CRSwNP) and without polyps (CRSsNP) were identified. Spatial modeling from pollutant monitoring sites was used to estimate exposures for patients meeting the inclusion criteria (total, n = 125; CRSsNP, n = 67; CRSsNP, n = 58). Skin-prick, intradermal dilutional, and in-vitro testing methods were utilized to determine aeroallergen sensitization. Disease severity indicators were measured by modified Lund-Mackay score (LMS), the 22-item Sino-Nasal Outcome Test (SNOT-22), systemic steroid therapy, and number of functional endoscopic sinus surgeries (FESS). RESULTS: Thirty-six percent (n = 45) of patients who described rhinitis symptoms demonstrated no reactivity to aeroallergen testing. Sixty-four percent (n = 80) tested positive for at least 1 allergen, with no differences found between CRSsNP and CRSwNP (62.1% vs 67.2%). There were significant differences in air pollutants between patients testing negative and positive for allergies (nonallergic vs allergic: PM2.5 , 11.32 vs 11.07 µg/m3 , p = 0.030; BC, 0.81 vs 0.76 absorbance, p =0.044). Nonallergic CRSwNP demonstrated higher PM2.5 compared with allergic counterparts (11.48 vs 11.09 µg/m3 , p = 0.032). A similar pattern was observed with BC (0.82 vs 0.75 absorbance, p = 0.017). In CRSsNP, BC correlated significantly with SNOT-22 (r = 0.55, p = 0.042). CONCLUSION: Our results suggest that small inhalant pollutants may contribute to nonallergic symptomatology in patients with and without nasal polyps. Regardless of allergy status, BC may play a role in CRS symptom severity.

Air pollutants may be environmental risk factors in chronic rhinosinusitis disease progression.

BACKGROUND: Little is known about the role of environmental exposures in the pathophysiology of chronic rhinosinusitis (CRS). In this study, we measured the impact of air pollutants (particulate matter 2.5 [PM2.5 ] and black carbon [BC]) on CRS with nasal polyposis (CRSwNP) and CRS without nasal polyposis (CRSsNP). METHODS: Spatial modeling from pollutant monitoring sites was used to estimate exposures surrounding residences for patients meeting inclusion criteria (total patients, n = 234; CRSsNP, n = 96; CRSwNP, n = 138). Disease severity outcome measures included modified Lund-Mackay score (LMS), systemic steroids, number of functional endoscopic sinus surgeries (FESS), and 22-item Sino-Nasal Outcome Test (SNOT-22) score. PM2.5 and BC exposures were correlated with outcome measures. RESULTS: Mean PM2.5 and BC findings were not significantly different between CRSwNP and CRSsNP patients or patients with and without asthma. Among those with CRSsNP, PM2.5 was significantly associated with undergoing FESS. For each unit increase in PM2.5 , there was a 1.89-fold increased risk in the proportion of CRSsNP patients who required further surgery (p = 0.015). This association was not identified in CRSwNP patients (p = 0.445). BC was also significantly associated with SNOT-22 score in the CRSsNP group. For each 0.1-unit increase in BC, there was a 7.97-unit increase in SNOT-22 (p = 0.008). A similar, although not significant, increase in SNOT-22 was found with increasing BC in the CRSwNP group (p = 0.728). CONCLUSION: Air pollutants correlate with CRS symptom severity that may be influenced by exposure levels, with a more pronounced impact on CRSsNP patients. This study is the first to demonstrate the possible role of inhalant pollutants in CRS phenotypes, addressing a critical knowledge gap in environmental risk factors for disease progression.

Cardiopulmonary exercise testing in the evaluation of liver disease in adults who have had the Fontan operation.

BACKGROUND: Liver disease (LD) is a long-term complication in patients with a single ventricle who have had the Fontan operation. A decline in cardiopulmonary exercise testing (CPET) variables is associated with increased risk of hospitalization, but its association with LD is unknown. AIM: To determine the association between CPET variables and LD in adults who have had the Fontan operation. METHODS: We retrospectively reviewed the medical records from two tertiary institutions. RESULTS: We identified 114 adults (≥18 years; mean 30.9±7.4 years) who had undergone the Fontan operation: 56% were women; 63% had total cavopulmonary connection; 66% had New York Heart Association (NYHA) class I status; 42% had arrhythmias; 22% had systemic right ventricle; and 35% had ventricular dysfunction. Of 81 patients with liver-imaging data, 41% had LD (i.e. imaging evidence of cirrhosis, with or without portal hypertension, splenomegaly or varices). There were no differences in clinical or echocardiographic variables between those with and without LD. Among the 58 patients with CPET data, mean peak oxygen consumption (VO2) was 18.6±5.7mL/kg/min, per-cent-predicted peak VO2 was 53.9±15.5%, peak oxygen pulse was 9.3±2.9mL/beat and per-cent-predicted peak oxygen pulse was 82.6±21.5%. Of the 44 patients with liver and CPET data, each standard deviation decrease in per-cent-predicted peak VO2 (16%) and per-cent-predicted peak oxygen pulse (22%) was associated with a 2.3-fold increase in the odds of LD, after adjusting for NYHA, institution and Fontan type (P=0.04). Similarly, each standard deviation decrease in per-cent-predicted peak VO2 and oxygen pulse was associated with an estimated 5.9-year and 4.9-year earlier onset of LD, respectively (P>0.05). CONCLUSIONS: Decline in per-cent-predicted peak VO2 and oxygen pulse was associated with increased odds of LD in adults who had undergone the Fontan operation. Our study supports more rapid hepatic evaluation among patients with abnormal or worsening CPET variables.

Aptamer-functionalized nanoparticles for surface immobilization-free electrochemical detection of cortisol in a microfluidic device.

Monitoring the periodic diurnal variations in cortisol from small volume samples of serum or saliva is of great interest, due to the regulatory role of cortisol within various physiological functions and stress symptoms. Current detection assays are immunologically based and require cumbersome antibody immobilization chemistries, thereby limiting the assay versatility, kinetics, and reproducibility. We present a quantitative aptamer-based detection methodology for cortisol that does not require target labeling, capture probe immobilization on the detection surface or wash steps prior to readout. Using a recognition system of aptamer functionalized gold nanoparticles pre-bound with electro-active triamcinolone, the cortisol level is detected based on its competitive binding to the aptamer by following signal from the displaced triamcinolone using square wave voltammetry at patterned graphene-modified electrodes in a microfluidic or nanoslit device. Due to the 3D analyte diffusion profile at the aptamer interface and the ability to enhance the surface area for cortisol capture, this assay shows signal linearity over a five-log analyte concentration range (10 µg/mL to 30 pg/mL) and exhibits rapid binding kinetics with cortisol versus other glucocorticoids, as apparent from the absence of interferences from estradiol, testosterone and progesterone. The assay is carried out within the biologically relevant range for glucocorticoids in serum and saliva matrices, and benchmarked versus ELISA and radioimmunoassays. Based on absence of cumbersome surface immobilization and wash steps for carrying out this assay, its quantitative signal characteristics and its ability to resist interferences from other glucocorticoids, we envision its application towards routine monitoring of cortisol within bio-fluids.

Validation of an 8-14 microm cloud monitor using visual observations of Antarctic cloud cover.

A cloud monitor instrument, sensing 8-14 microm infrared radiation, has been installed near McMurdo, Antarctica, in order to provide supporting cloud cover information for a high-resolution sky-viewing Fabry-Perot spectrometer experiment. Validation of the cloud monitor observations was performed by a 1 year long comparison with the available 6 h visual cloud observations, expressed in octas (obscured eighths of the sky). The agreement between visual observations and those of the cloud monitor is good--the variation being slightly over one step, or octa--except at the clear sky extreme, which the subjective visual observations tend to overestimate during dark and moonless conditions. Both visual observations and the cloud monitor measurements show an austral winter-summer difference with the winter being clearer.

Two-generation reproduction and cross-foster studies of perfluorooctanesulfonate (PFOS) in rats.

Perfluorooctanesulfonate (PFOS) is a persistent acid found widely distributed in wildlife and humans. To understand the potential reproductive and developmental effects of PFOS, a two-generation reproduction study was conducted in rats. Male and female rats were dosed via oral gavage at dose levels of 0, 0.1, 0.4, 1.6, and 3.2 mg/(kg day) for 6 weeks prior to mating, during mating, and, for females, through gestation and lactation, across two generations. Due to substantial F1 neonatal toxicity observed in the 1.6 and 3.2 mg/(kg day) groups, continuation into the second generation was limited to F1 pups from the 0, 0.1, and 0.4 mg/(kg day) groups. No adverse effects were observed in F0 females or their fetuses upon caesarean sectioning at gestation day 10. Statistically significant reductions in body-weight gain and feed consumption were observed in F0 generation males and females at dose levels of 0.4 mg/(kg day) and higher, but not in F1 adults. PFOS did not affect reproductive performance (mating, estrous cycling, and fertility); however, reproductive outcome, as demonstrated by decreased length of gestation, number of implantation sites, and increased numbers of dams with stillborn pups or with all pups dying on lactation days 1-4, was affected at 3.2 mg/(kg day) in F0 dams. These effects were not observed in F1 dams at the highest dose tested, 0.4 mg/(kg day). Neonatal toxicity in F1 pups, as demonstrated by reduced survival and body-weight gain through the end of lactation, occurred at a maternal dose of 1.6 mg/(kg day) and higher while not at dose levels of 0.1 or 0.4 mg/(kg day) or in F2 pups at the 0.1 or 0.4 mg/(kg day) dose levels tested. In addition to these adverse effects, slight yet statistically significant developmental delays occurred at 0.4 (eye opening) and 1.6 mg/(kg day) (eye opening, air righting, surface righting, and pinna unfolding) in F1 pups. Based on these data, the NOAELs were as follows: reproductive function: F0> or =3.2 and F1> or =0.4 mg/(kg day); reproductive outcome: F0=1.6 and F1> or =0.4 mg/(kg day); overall parental effects: F0=0.1 and F1> or =0.4 mg/(kg day); offspring effects: F0=0.4 and F1> or =0.4 mg/(kg day). To distinguish between maternal and pup influences contributing to the perinatal mortality observed in the two-generation study, a follow-up cross-foster study was performed. Results of this study indicated that in utero exposure to PFOS causally contributed to post-natal pup mortality, and that pre-natal and post-natal exposure to PFOS was additive with respect to the toxic effects observed in pups.