Editing Myosin VB Gene to Create Porcine Model of Microvillus Inclusion Disease, With Microvillus-Lined Inclusions and Alterations in Sodium Transporters.

Gastroenterology. 2020 06;158(8):2236-2249.e9

Authors: Engevik AC, Coutts AW, Kaji I, Rodriguez P, Ongaratto F, Saqui-Salces M, Medida RL, Meyer AR, Kolobova E, Engevik MA, Williams JA, Shub MD, Carlson DF, Melkamu T, Goldenring JR

Abstract
BACKGROUND & AIMS: Microvillus inclusion disease (MVID) is caused by inactivating mutations in the myosin VB gene (MYO5B). MVID is a complex disorder characterized by chronic, watery, life-threatening diarrhea that usually begins in the first hours to days of life. We developed a large animal model of MVID to better understand its pathophysiology.
METHODS: Pigs were cloned by transfer of chromatin from swine primary fetal fibroblasts, which were edited with TALENs and single-strand oligonucleotide to introduce a P663-L663 substitution in the endogenous swine MYO5B (corresponding to the P660L mutation in human MYO5B, associated with MVID) to fertilized oocytes. We analyzed duodenal tissues from patients with MVID (with the MYO5B P660L mutation) and without (controls), and from pigs using immunohistochemistry. Enteroids were generated from pigs with MYO5B(P663L) and without the substitution (control pigs).
RESULTS: Duodenal tissues from patients with MVID lacked MYO5B at the base of the apical membrane of intestinal cells; instead MYO5B was intracellular. Intestinal tissues and derived enteroids from MYO5B(P663L) piglets had reduced apical levels and diffuse subapical levels of sodium hydrogen exchanger 3 and SGLT1, which regulate transport of sodium, glucose, and water, compared with tissues from control piglets. However, intestinal tissues and derived enteroids from MYO5B(P663L) piglets maintained CFTR on apical membranes, like tissues from control pigs. Liver tissues from MYO5B(P663L) piglets had alterations in bile salt export pump, a transporter that facilitates bile flow, which is normally expressed in the bile canaliculi in the liver.
CONCLUSIONS: We developed a large animal model of MVID that has many features of the human disease. Studies of this model could provide information about the functions of MYO5B and MVID pathogenesis, and might lead to new treatments.

PMID: 32112796 [PubMed - indexed for MEDLINE]

Klin Padiatr. 2021 Feb 18. doi: 10.1055/a-1341-1698. Online ahead of print.

ABSTRACT

BACKGROUND: Cystic fibrosis (CF) is a multisystemic disease that prevalently involves the lungs. Hypoxemia occurs due to the existing of progressive damage to the pulmonary parenchyma and pulmonary vessels. The condition may cause systolic and diastolic dysfunction to the right ventricle due to the effects of high pulmonary artery systolic pressure (PASP). The study aimed to determine echocardiographic alterations in PASP, right ventricle (RV) anatomy, and functions in mild CF children.

MATERIALS AND METHODS: RV anatomy, systolic, and diastolic functions were evaluated with conventional echocardiographic measurements. Estimated PASP was used measured with new echocardiographic modalities, including pulmonary artery acceleration time (PAAT), right ventricular ejection time (RVET), and their ratio (PAAT/RVET). The obtained echocardiographic data were statistically compared between the patient group and the control group.

RESULTS: The study consisted of 30 pediatric patients with mild CF and 30 healthy children with similar demographics. In patient group, conventional parameters disclosed differences in RV anatomy, both systolic and diastolic functions of RV compared with the healthy group. We did not compare the patient group with published standard data because of the wide range variability. However, new echocardiographic parameters showed notable increase in pulmonary artery pressure compared with values of control group and published standard data (p<0.001).

CONCLUSION: Elevated PASP, RV failure, and Cor pulmonale usually begin early in children with mild CF. In addition to routine echocardiographic measurements to evaluate RV, we recommend the use of new echocardiographic modalities for routine examinations and in the follow up of children with mild CF.

PMID:33601431 | DOI:10.1055/a-1341-1698

Ann Am Thorac Soc. 2021 Feb 18. doi: 10.1513/AnnalsATS.202008-1054OC. Online ahead of print.

ABSTRACT

RATIONALE: Previous studies showed that lumacaftor-ivacaftor therapy results in partial rescue of cystic fibrosis transmembrane conductance regulator (CFTR) activity and moderate improvement of spirometry in Phe508del homozygous patients with cystic fibrosis (CF). However, the effects of lumacaftor-ivacaftor on lung clearance index (LCI), lung morphology and perfusion detected by chest magnetic resonance imaging (MRI), and effects on the airway microbiome and inflammation remain unknown.

OBJECTIVES: To investigate the effects of lumacaftor-ivacaftor on LCI, lung MRI scores, and airway microbiome and inflammation.

METHODS: In this prospective observational study we assessed clinical outcomes including spirometry and body mass index, LCI, lung MRI scores, sputum microbiome and pro-inflammatory cytokines in 30 Phe508del homozygous patients with CF 12 years and older before and 8-16 weeks after initiation of lumacaftor-ivacaftor therapy.

MEASUREMENTS AND MAIN RESULTS: Lumacaftor-ivacaftor had no effects on FEV1 % predicted (1.7%, 95% confidence interval (CI) -1.0 to 4.3%; P = 0.211), but improved LCI (-1.6, 95% CI -2.6 to -0.5; P < 0.01) and MRI morphology (-1.3, 95% CI -2.3 to -0.3; P < 0.05) and perfusion score (-1.2, 95% CI -2.3 to -0.2; P < 0.05) in our study cohort. Further, lumacaftor-ivacaftor decreased the total bacterial load (-1.8, 95% CI -3.3 to -0.34; P < 0.05) and increased the Shannon diversity of the airway microbiome (0.4, 95% CI 0.1 to 0.8; P < 0.05), and reduced IL-1β levels (median change -324.2 pg/ml, 95% CI -938.7 to 290.4 pg/ml; P < 0.05) in sputum of Phe508del homozygous patients.

CONCLUSIONS: This study shows that lumacaftor-ivacaftor has beneficial effects on lung ventilation, morphology and perfusion, as well as on the airway microbiome and inflammation in Phe508del homozygous patients. Our results suggest that LCI and MRI may be more sensitive than FEV1 % predicted to detect response to CFTR modulator therapy in patients with chronic CF lung disease. Clinical trial registered with ClinicalTrials.gov (NCT02807415).

PMID:33600745 | DOI:10.1513/AnnalsATS.202008-1054OC

Am J Respir Crit Care Med. 2021 Feb 18. doi: 10.1164/rccm.202011-4153OC. Online ahead of print.

ABSTRACT

RATIONALE: Elexacaftor-tezacaftor-ivacaftor is a cystic fibrosis transmembrane conductance regulator modulator combination, developed for cystic fibrosis (CF) patients with at least one Phe508del mutation.

OBJECTIVES: To evaluate the effects of elexacaftor-tezacaftor-ivacaftor in CF patients with advanced respiratory disease.

METHODS: A prospective observational study, including all patients aged ≥12 years and with ppFEV1<40 who initiated elexacaftor-tezacaftor-ivacaftor from December 2019 to August 2020 in France was conducted. Clinical characteristics were collected at initiation and at one and three months. Safety and effectiveness were evaluated by September 2020. National-level transplantation and mortality figures for 2020 were obtained from the French CF and transplant centers and registries.

MAIN RESULTS: Elexacaftor-tezacaftor-ivacaftor was initiated in 245 patients with a median [IQR] ppFEV1=29 [24; 34]. Mean (95% confidence interval) absolute increase in ppFEV1 was +15.1 (+13.8,+16.4; P<0.0001) and in weight was +4.2 kg (+3.9, +4.6; P<0.0001). The number of patients requiring long-term oxygen, non-invasive ventilation and/or enteral tube feeding decreased by 50%, 30% and 50%, respectively (P<0.01). Although 16 patients were on the transplant waiting list and 37 were undergoing transplantation evaluation at treatment initiation, only two were transplanted and one died. By September 2020 only five patients were still on transplantation path. Compared with the previous two years, a two-fold decrease in the number of lung transplantations in CF patients was observed in 2020, while the number of death without transplantation remained stable.

CONCLUSIONS: In patients with advanced disease, elexacaftor-tezacaftor-ivacaftor is associated with rapid clinical improvement, often leading to the indication for lung transplantation being suspended.

PMID:33600738 | DOI:10.1164/rccm.202011-4153OC

J Pediatr Endocrinol Metab. 2020 Oct 22;21(6):605. doi: 10.1515/jpem-2008-210617.

NO ABSTRACT

PMID:33600695 | DOI:10.1515/jpem-2008-210617

Microb Genom. 2021 Feb 18. doi: 10.1099/mgen.0.000510. Online ahead of print.

ABSTRACT

Staphylococcus aureus chronic airway infection in patients with cystic fibrosis (CF) allows this pathogen to adapt over time in response to different selection pressures. We have previously shown that the main sequence types related to community-acquired methicillin-resistant S. aureus (MRSA) infections in Argentina - ST5 and ST30 - are also frequently isolated from the sputum of patients with CF, but in these patients they usually display multi-drug antimicrobial resistance. In this study, we sequenced the genomes of MRSA from four paediatric CF patients with the goal of identifying mutations among sequential isolates, especially those possibly related to antimicrobial resistance and virulence, which might contribute to the adaptation of the pathogen in the airways of patients with CF. Our results revealed genetic differences in sequential MRSA strains isolated from patients with CF in both their core and accessory genomes. Although the genetic adaptation of S. aureus was distinct in different hosts, we detected independent mutations in thyA, htrA, rpsJ and gyrA - which are known to have crucial roles in S. aureus virulence and antimicrobial resistance - in isolates recovered from multiple patients. Moreover, we identified allelic variants that were detected in all of the isolates recovered after a certain time point; these non-synonymous mutations were in genes associated with antimicrobial resistance, virulence, iron scavenging and oxidative stress resistance. In conclusion, our results provide evidence of genetic variability among sequential MRSA isolates that could be implicated in the adaptation of these strains during chronic CF airway infection.

PMID:33599606 | DOI:10.1099/mgen.0.000510

mSphere. 2021 Feb 17;6(1):e01250-20. doi: 10.1128/mSphere.01250-20.

ABSTRACT

Aspergillus fumigatus is a filamentous fungus which can cause multiple diseases in humans. Allergic broncho-pulmonary aspergillosis (ABPA) is a disease diagnosed primarily in cystic fibrosis patients caused by a severe allergic response often to long-term A. fumigatus colonization in the lungs. Mice develop an allergic response to repeated inhalation of A. fumigatus spores; however, no strains have been identified that can survive long-term in the mouse lung and cause ABPA-like disease. We characterized A. fumigatus strain W72310, which was isolated from the expectorated sputum of an ABPA patient, by whole-genome sequencing and in vitro and in vivo viability assays in comparison to a common reference strain, CEA10. W72310 was resistant to leukocyte-mediated killing and persisted in the mouse lung longer than CEA10, a phenotype that correlated with greater resistance to oxidative stressors, hydrogen peroxide, and menadione, in vitro In animals both sensitized and challenged with W72310, conidia, but not hyphae, were viable in the lungs for up to 21 days in association with eosinophilic airway inflammation, airway leakage, serum IgE, and mucus production. W72310-sensitized mice that were recall challenged with conidia had increased inflammation, Th1 and Th2 cytokines, and airway leakage compared to controls. Collectively, our studies demonstrate that a unique strain of A. fumigatus resistant to leukocyte killing can persist in the mouse lung in conidial form and elicit features of ABPA-like disease.IMPORTANCE Allergic broncho-pulmonary aspergillosis (ABPA) patients often present with long-term colonization of Aspergillus fumigatus Current understanding of ABPA pathogenesis has been complicated by a lack of long-term in vivo fungal persistence models. We have identified a clinical isolate of A. fumigatus, W72310, which persists in the murine lung and causes an ABPA-like disease phenotype. Surprisingly, while viable, W72310 showed little to no growth beyond the conidial stage in the lung. This indicates that it is possible that A. fumigatus can cause allergic disease in the lung without any significant hyphal growth. The identification of this strain of A. fumigatus can be used not only to better understand disease pathogenesis of ABPA and potential antifungal treatments but also to identify features of fungal strains that drive long-term fungal persistence in the lung. Consequently, these observations are a step toward helping resolve the long-standing question of when to utilize antifungal therapies in patients with ABPA and fungal allergic-type diseases.

PMID:33597172 | DOI:10.1128/mSphere.01250-20

Eur Respir Rev. 2021 Feb 16;30(159):200293. doi: 10.1183/16000617.0293-2020. Print 2021 Mar 31.

ABSTRACT

The development of cystic fibrosis-related diabetes (CFRD) often leads to poorer outcomes in patients with cystic fibrosis including increases in pulmonary exacerbations, poorer lung function and early mortality. This review highlights the many factors contributing to the clinical decline seen in patients diagnosed with CFRD, highlighting the important role of nutrition, the direct effect of hyperglycaemia on the lungs, the immunomodulatory effects of high glucose levels and the potential role of genetic modifiers in CFRD.

PMID:33597125 | DOI:10.1183/16000617.0293-2020

Reproductive Health Counseling and Contraceptive Use in Adolescents with Cystic Fibrosis.

Pediatr Pulmonol. 2021 Feb 16;:

Authors: Hernandez AM, Burdick B, Adeyemi-Fowode

Abstract
BACKGROUND: Cystic Fibrosis (CF) is a progressive, genetic disease posing reproductive health concerns to affected women such as high-risk pregnancies and medication interactions leading to contraceptive failure. Reproductive health counseling in this population is of the utmost importance but barriers to providing it include lack of time, knowledge, and provider discomfort. We sought to evaluate reproductive health counseling and contraceptive use in female adolescent CF patients.
METHODS: An IRB approved retrospective chart review was performed between March 2008 and March 2018. Females 10-21 years old with the diagnosis of CF were reviewed. Descriptive statistics were used.
RESULTS: Thirty-three patients met inclusion criteria: 16 non-sexually active and 17 sexually active. Thirteen patients were counseled about pregnancy risks. All sexually active patients used contraception with the most common being condoms and combined oral contraceptive pills. Six pregnancies occurred resulting in five live births and one termination.
CONCLUSIONS: Less than half of patients were counseled about pregnancy and contraception and most patients chose high failure methods. Female adolescents with CF should be routinely screened for sexual activity, offered long acting reversible contraception (LARC), and counseled on the effects of CF on pregnancy. This article is protected by copyright. All rights reserved.

PMID: 33590969 [PubMed - as supplied by publisher]

Disparities in insurance coverage among hospitalized adult congenital heart disease patients before and after the Affordable Care Act.

Birth Defects Res. 2021 Feb 15;:

Authors: Salciccioli KB, Salemi JL, Broda CR, Lopez KN

Abstract
BACKGROUND: Data are lacking regarding the insurance status of adults with congenital heart disease (ACHD). We investigated whether the Affordable Care Act (ACA) impacted insurance status among hospitalized ACHD, identified associated sociodemographic factors, and compared coverage to adults with other chronic childhood conditions.
METHODS: Serial cross-sectional analysis of National Inpatient Sample hospitalizations from 2007 to 2016 was performed for patients 18-64 years old. ACHD were identified using ICD-9/10-CM codes and compared to patients with sickle cell disease (SCD), cystic fibrosis (CF), and the general population. Age was dichotomized as 18-25 years (transition aged) or 26-64 years. Groups were compared by era (pre-ACA [January 2007-June 2010]; early-ACA [July 2010-December 2013], which eliminated pre-existing condition exclusions; and full-ACA [January 2014-December 2016]) using interrupted time series and multivariable Poisson regression analyses.
RESULTS: Overall, uninsured hospitalizations decreased from pre-ACA (12.0%) to full-ACA (8.5%). After full ACA implementation, ACHD had lower uninsured rates than the general hospitalized population (6.0 vs. 8.6%, p < .01), but higher rates than those with other chronic childhood diseases (SCD [4.5%]; CF [1.6%]). Across ACA eras, transition aged ACHD had higher uninsured rates than older patients (8.9 vs. 7.6%, p < .01), and Hispanic patients remained less insured than other groups.
CONCLUSIONS: Hospitalized ACHD were better insured than the general population but less insured than those with SCD or CF. Full ACA implementation was associated with improved insurance coverage for all groups, but disparities persisted for transition aged and Hispanic patients. Ongoing evaluation of the effects of insurance and health policy on ACHD remains critical to diminish health disparities.

PMID: 33590705 [PubMed - as supplied by publisher]

A novel highly bio-available itraconazole formulation (SUBA®-Itraconazole) for anti-fungal prophylaxis in Lung Transplant recipients.

Transpl Infect Dis. 2021 Feb 16;:e13587

Authors: Whitmore TJ, Yaw M, Lavender M, Musk M, Boan P, Wrobel J

Abstract
Antifungal prophylaxis remains a mainstay of lung transplantation, given invasive fungal infection is a common and serious complication after lung transplantation. Choice of systemic agent to prevent invasive fungal infection varies between centres and funding of agents remains challenging. Our centre has recently changed from posaconazole to a highly bioavailable formulation of itraconazole (SUBA®-itraconazole) at substantially reduced cost, but safety and toxicity require further assessment. A retrospective study of lung transplant patients receiving systemic antifungal prophylaxis from December 2016 through December 2019 following change from posaconazole to itraconazole as standard practice. 150 patients with lung transplants were managed in this time period, with 88 (59%) receiving at least 1 mold-active triazole during the study period. 48 (58%) of these patients received SUBA®-itraconazole; 68 (82%) received posaconazole and 10 (12%) received voriconazole. The average cost per patient during the study period was significantly lower on SUBA®-itraconazole (mean $1,548/patient/6 month course) than posaconazole (mean $16,921.35/patient/6 month course). Target trough concentrations for prophylaxis of itraconazole >0.5 mg/L and posaconazole >0.7 mg/L were achieved on empiric dosing in 49% and 68% respectively. Overall trough itraconazole (0.50 vs 1.12 mg/L, p < 0.001) and posaconazole (1.37 vs 2.10 mg/L p < 0.001) concentrations were significantly lower in patients with cystic fibrosis. Calcineurin inhibitor dose changes on introduction or cessation were similar for SUBA®-itraconazole and posaconazole. Breakthrough invasive fungal infection and toxicity were rare. SUBA®-itraconazole is well tolerated, associated with rare breakthrough invasive fungal infection, and lower cost. Prospective studies following general introduction are required to determine long-term safety, tolerability and efficacy.

PMID: 33590676 [PubMed - as supplied by publisher]

How Many Maneuvers Should We Do for Maximal Inspiratory and Expiratory Muscle Pressure Testing in Children: A Retrospective Review in Children with Cystic Fibrosis.

Lung. 2021 Feb 15;:

Authors: Boonjindasup W, Chang AB, Marchant JM, Irons JY, McElrea MS

Abstract
OBJECTIVES: Maximal inspiratory pressure (MIP) and maximal expiratory pressure (MEP) could be useful clinical parameters in monitoring many conditions including cystic fibrosis (CF). However, current protocols for undertaking the measurements lack standardization including the number of repeated attempts to achieve best values. We aimed to (a) determine the optimum number of attempts to achieve best MIP/MEP values, and (b) evaluate if the number of attempts is consistent across two different test days.
METHODS: We analyzed data of a previous randomized controlled trial involving the effect of singing on respiratory muscle strength in 35 children with CF. On two different days (T1, T2) children performed MIP/MEP with at least ten attempts each to achieve < 10% repeatability.
RESULTS: All children achieved repeatable MIP/MEP values within 10-11 attempts with 24 (68.6%) and 26 (74.3%) of these achieving best values of MIP and MEP, respectively, at attempts 6-11. Median values of the pressures by three, five, eight and all attempts significantly increased with more attempts (all p < 0.05). At T2, 56% required fewer attempts to achieve best values, but 32% required more attempts, indicating that the number of attempts required was inconsistent between test days.
CONCLUSION: It is likely that at least ten attempts (best two within < 10% variability) is required to achieve best and reliable MIP/MEP in children with CF. A larger sample size in children with CF and various conditions is required to consolidate these findings.

PMID: 33590270 [PubMed - as supplied by publisher]

Humoral signatures of protective and pathological SARS-CoV-2 infection in children.

Nat Med. 2021 Feb 12;:

Authors: Bartsch YC, Wang C, Zohar T, Fischinger S, Atyeo C, Burke JS, Kang J, Edlow AG, Fasano A, Baden LR, Nilles EJ, Woolley AE, Karlson EW, Hopke AR, Irimia D, Fischer ES, Ryan ET, Charles RC, Julg BD, Lauffenburger DA, Yonker LM, Alter G

Abstract
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic continues to spread relentlessly, associated with a high frequency of respiratory failure and mortality. Children experience largely asymptomatic disease, with rare reports of multisystem inflammatory syndrome in children (MIS-C). Identifying immune mechanisms that result in these disparate clinical phenotypes in children could provide critical insights into coronavirus disease 2019 (COVID-19) pathogenesis. Using systems serology, in this study we observed in 25 children with acute mild COVID-19 a functional phagocyte and complement-activating IgG response to SARS-CoV-2, similar to the acute responses generated in adults with mild disease. Conversely, IgA and neutrophil responses were significantly expanded in adults with severe disease. Moreover, weeks after the resolution of SARS-CoV-2 infection, children who develop MIS-C maintained highly inflammatory monocyte-activating SARS-CoV-2 IgG antibodies, distinguishable from acute disease in children but with antibody levels similar to those in convalescent adults. Collectively, these data provide unique insights into the potential mechanisms of IgG and IgA that might underlie differential disease severity as well as unexpected complications in children infected with SARS-CoV-2.

PMID: 33589825 [PubMed - as supplied by publisher]

Contribution of common CFTR variants (M470V, T854, and Q1463) to cystic fibrosis in Tunisia: haplotype analysis.

Ann Biol Clin (Paris). 2021 Feb 15;:

Authors: Nefzi M, Fredj SH, Dabboubi R, Hamouda S, Tebib N, Boussetta K, Messaoud T

Abstract
BACKGROUND & OBJECTIVES: Cystic fibrosis (CF) is caused by mutations in the gene encoding the CF transmembrane regulator (CFTR) protein, a chloride channel located in the epithelial cell membrane. Over than 2,000 CFTR mutations have been identified, which contribute to the variety of clinical phenotypes of CF. We performed a case-control study to determine p.Met470Val (M470V), p.Thr854= (T854) and p.Gln1463= (Q1463) polymorphisms frequencies in CF patients and healthy controls and to elaborate haplotype based on these SNPs.
METHODS: The genotyping of M470V (exon 10), T854 (exon 14a), and Q1463 (exon 24) variants were identified using polymorphism restriction fragment length polymorphism (RFLP).
RESULTS & CONCLUSION: Statistical difference was noted in the genotype distribution of two markers, M470V and T854, between CF and control groups. However, the Q1463 polymorphism is not identified in two studied groups. Three haplotypes were found in CF patients and controls. An exclusive association between the ancestral haplotype 1-1-2 and p.Phe508del (F508del) mutation was shown. In Tunisia, this is the first work to be interested in the analysis of M470V, T854 and Q1463 polymorphisms and haplotypes associated with the most common mutation, F508del, in the Tunisian population and worldwide.

PMID: 33589413 [PubMed - as supplied by publisher]

Neutrophil dysfunction in cystic fibrosis.

J Cyst Fibros. 2021 Feb 12;:

Authors: Yonker LM, Marand A, Muldur S, Hopke A, Leung HM, De La Flor D, Park G, Pinsky H, Guthrie LB, Tearney GJ, Irimia D, Hurley BP

Abstract
BACKGROUND: Excessive neutrophil inflammation is the hallmark of cystic fibrosis (CF) airway disease. Novel technologies for characterizing neutrophil dysfunction may provide insight into the nature of these abnormalities, revealing a greater mechanistic understanding and new avenues for CF therapies that target these mechanisms.
METHODS: Blood was collected from individuals with CF in the outpatient clinic, CF individuals hospitalized for a pulmonary exacerbation, and non-CF controls. Using microfluidic assays and advanced imaging technologies, we characterized 1) spontaneous neutrophil migration using microfluidic motility mazes, 2) neutrophil migration to and phagocytosis of Staphylococcal aureus particles in a microfluidic arena, 3) neutrophil swarming on Candida albicans clusters, and 4) Pseudomonas aeruginosa-induced neutrophil transepithelial migration using micro-optical coherence technology (µOCT).
RESULTS: Participants included 44 individuals: 16 Outpatient CF, 13 Hospitalized CF, and 15 Non-CF individuals. While no differences were seen with spontaneous migration, CF neutrophils migrated towards S. aureus particles more quickly than non-CF neutrophils (p < 0.05). CF neutrophils, especially Hospitalized CF neutrophils, generated significantly larger aggregates around S. aureus particles over time. Hospitalized CF neutrophils were more likely to have dysfunctional swarming (p < 0.01) and less efficient clearing of C. albicans (p < 0.0001). When comparing trans-epithelial migration towards Pseudomonas aeruginosa epithelial infection, Outpatient CF neutrophils displayed an increase in the magnitude of transmigration and adherence to the epithelium (p < 0.05).
CONCLUSIONS: Advanced technologies for characterizing CF neutrophil function reveal significantly altered migratory responses, cell-to-cell clustering, and microbe containment. Future investigations will probe mechanistic basis for abnormal responses in CF to identify potential avenues for novel anti-inflammatory therapeutics.

PMID: 33589340 [PubMed - as supplied by publisher]

Burkholderia vietnamiensis Causing a Non-lactational Breast Abscess in a Non-cystic Fibrosis Patient in Tamil Nadu, India.

Indian J Med Microbiol. 2020 Jul-Dec;38(3-4):496-499

Authors: Rohit A, Rani MS, Anand NS, Chellappa C, Mohanapriya P, Karunasagar I, Karunasagar I, Deekshit VK

Abstract
Burkholderia cepacia complex is a Gram-negative opportunistic pathogen usually found in people with an immunocompromised condition such as cystic fibrosis (CF). In a tropical country like India, this organism has been associated with a number of hospital-acquired infections including sepsis. We present here a report of a case of Burkholderia vietnamiensis causing a non-lactational breast abscess in a non-CF patient. The pathogen was identified as B. cepacia using Vitek system and matrix-assisted laser desorption ionisation-time of flight. This was confirmed by polymerase chain reaction (PCR) using recA genus-specific gene and sequencing of the PCR amplicons. recA-restriction fragment length polymorphism and recA gene sequencing revealed that the isolate is B. vietnamiensis. This is the first description of B. vietnamiensis isolated from a clinical case from India.

PMID: 33589214 [PubMed - in process]

GRAF2, WDR44, and MICAL1 mediate Rab8/10/11-dependent export of E-cadherin, MMP14, and CFTR ΔF508.

J Cell Biol. 2020 05 04;219(5):

Authors: Lucken-Ardjomande Häsler S, Vallis Y, Pasche M, McMahon HT

Abstract
In addition to the classical pathway of secretion, some transmembrane proteins reach the plasma membrane through alternative routes. Several proteins transit through endosomes and are exported in a Rab8-, Rab10-, and/or Rab11-dependent manner. GRAFs are membrane-binding proteins associated with tubules and vesicles. We found extensive colocalization of GRAF1b/2 with Rab8a/b and partial with Rab10. We identified MICAL1 and WDR44 as direct GRAF-binding partners. MICAL1 links GRAF1b/2 to Rab8a/b and Rab10, and WDR44 binds Rab11. Endogenous WDR44 labels a subset of tubular endosomes, which are closely aligned with the ER via binding to VAPA/B. With its BAR domain, GRAF2 can tubulate membranes, and in its absence WDR44 tubules are not observed. We show that GRAF2 and WDR44 are essential for the export of neosynthesized E-cadherin, MMP14, and CFTR ΔF508, three proteins whose exocytosis is sensitive to ER stress. Overexpression of dominant negative mutants of GRAF1/2, WDR44, and MICAL1 also interferes with it, facilitating future studies of Rab8/10/11-dependent exocytic pathways of central importance in biology.

PMID: 32344433 [PubMed - indexed for MEDLINE]

Intracellular Cl- Regulation of Ciliary Beating in Ciliated Human Nasal Epithelial Cells: Frequency and Distance of Ciliary Beating Observed by High-Speed Video Microscopy.

Int J Mol Sci. 2020 Jun 05;21(11):

Authors: Yasuda M, Inui TA, Hirano S, Asano S, Okazaki T, Inui T, Marunaka Y, Nakahari T

Abstract
Small inhaled particles, which are entrapped by the mucous layer that is maintained by mucous secretion via mucin exocytosis and fluid secretion, are removed from the nasal cavity by beating cilia. The functional activities of beating cilia are assessed by their frequency and the amplitude. Nasal ciliary beating is controlled by intracellular ions (Ca2+, H+ and Cl-), and is enhanced by a decreased concentration of intracellular Cl- ([Cl-]i) in ciliated human nasal epithelial cells (cHNECs) in primary culture, which increases the ciliary beat amplitude. A novel method to measure both ciliary beat frequency (CBF) and ciliary beat distance (CBD, an index of ciliary beat amplitude) in cHNECs has been developed using high-speed video microscopy, which revealed that a decrease in [Cl-]i increased CBD, but not CBF, and an increase in [Cl-]i decreased both CBD and CBF. Thus, [Cl-]i inhibits ciliary beating in cHNECs, suggesting that axonemal structures controlling CBD and CBF may have Cl- sensors and be regulated by [Cl-]i. These observations indicate that the activation of Cl- secretion stimulates ciliary beating (increased CBD) mediated via a decrease in [Cl-]i in cHNECs. Thus, [Cl-]i is critical for controlling ciliary beating in cHNECs. This review introduces the concept of Cl- regulation of ciliary beating in cHNECs.

PMID: 32517062 [PubMed - in process]

ERJ Open Res. 2021 Feb 8;7(1):00712-2020. doi: 10.1183/23120541.00712-2020. eCollection 2021 Jan.

ABSTRACT

The American Thoracic Society and European Respiratory Society commissioned a task force to update the technical standards for spirometry testing with the aim of increasing the accuracy, precision and quality of spirometry measurements and improving the patient experience. To inform the task force with patient experiences, the European Lung Foundation, in collaboration with the task force, conducted an online survey in 10 languages between August and September 2018. There were 1760 respondents from 52 countries. The majority were adults (97.1%); the most common reasons for spirometry referral were diagnosis (35.0%) and management of an ongoing condition (60.1%). 53.2% reported regularly using inhalers. Respondents were very experienced with spirometry: 89.9% completed more than one test; 48% completed 10 or more tests. However, most reported not knowing what forced expiratory volume in 1 s (FEV1) means (59.4%) and only 39.6% knew their most recent FEV1; the exception was respondents with cystic fibrosis who reported much greater knowledge. Respondents rated as moderately or seriously problematic: being told to keep blowing when they felt nothing is coming out (31.4%), coughing (30.4%), tiredness (26.3%) and concern about shortness of breath (20.1%). Overall, respondents found spirometry to be acceptable; however, an important minority (17%) found it difficult. Patients want clear information before, during and after the test, including information on stopping medications. Operators have an important role in increasing the ease of patients and changes to the testing environment can increase patient comfort. Patients want access to their results and want to understand how they relate to their individual health.

PMID:33585650 | PMC:PMC7869590 | DOI:10.1183/23120541.00712-2020

Front Cell Infect Microbiol. 2021 Jan 11;10:601821. doi: 10.3389/fcimb.2020.601821. eCollection 2020.

ABSTRACT

Aspergillus fumigatus (Af) frequently colonizes the respiratory tract of patients with cystic fibrosis (CF). Af is associated with loss of pulmonary function and allergic bronchopulmonary aspergillosis (ABPA), a hypersensitivity fungal lung disease. Environmental factors have impact on CF patients' lung function variation. The aim of this nationwide questionnaire survey was to investigate the amount of CF patients with frequent pet contact including pet species and to examine the potential impact of frequent pet contact on the occurrence of Af colonization and ABPA diagnosis in these patients. The survey was carried out in 31 German CF centers in 2018. A total of 1232 who completed the surveys were included, and statistical analysis was performed by chi-squared test. Within the study cohort 49.8% of subjects (n = 614; CF patients < 18years: 49.4%, n = 234; ≥ 18years: 50.1%, n = 380) reported frequent contact to pets, of which 60.7% reported frequent contact to dogs, 42.3% to cats and other animals. Of those with frequent pet contact, 71.8% (n = 441) had contact to one pet or more pets from the same family. Af colonization was not significantly associated with frequent pet contact. ABPA diagnosis was documented in 16.7% (n = 206) of all included CF patients and was significantly associated with frequent pet contact (18.9%, n = 116, p = 0.042), confirming previous single center examinations. Particularly, patients with frequent contact to dogs showed an increased ABPA prevalence of 21.3%. Frequent pet contact might be a risk factor for ABPA. CF patients who are sensitized to Af should be informed about the increased risk to develop an ABPA by frequent pet contact. Patients with recurrent onset of ABPA should be evaluated in terms of frequent pet contact.

PMID:33585274 | PMC:PMC7873990 | DOI:10.3389/fcimb.2020.601821