41 results on '"Nam Soo Joo"'
Search Results
2. Critical role of down-regulated in adenoma bicarbonate transporter in linaclotide stimulated intestinal bicarbonate secretion
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Jessica B. Sarthi, Annie M. Trumbull, Shayda M. Abazari, Vincent van Unen, Joshua E. Chan, Nam Soo Joo, Yanfen Jiang, Calvin J. Kuo, and Zachary M. Sellers
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Article - Abstract
Duodenal bicarbonate secretion is critical to epithelial protection, nutrient digestion/absorption and is impaired in cystic fibrosis (CF). We examined if linaclotide, typically used to treat constipation, may also alter duodenal bicarbonate secretion. Bicarbonate secretion was measured in vivo and in vitro using mouse and human duodenum. Ion transporter localization was identified with confocal microscopy and de novo analysis of human duodenal single cell RNA sequencing (sc-RNAseq) was performed. Linaclotide increased bicarbonate secretion in mouse and human duodenum in the absence of CFTR expression or function. Linaclotide-stimulated bicarbonate secretion was eliminated by down-regulated in adenoma (DRA) inhibition, regardless of CFTR activity. Sc-RNAseq identified that 70% of villus cells expressedSLC26A3, but notCFTR, mRNA. Linaclotide increased apical membrane expression of DRA in non-CF and CF differentiated enteroids. These data provide insights into the action of linaclotide and suggest linaclotide may be a useful therapy for CF individuals with impaired bicarbonate secretion.
- Published
- 2023
3. Combined agonists act synergistically to increase mucociliary clearance in a cystic fibrosis airway model
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Hyung Ju Cho, Meagan Shinbashi, John F. Engelhardt, Jae Young Choi, Jeffrey J. Wine, Nam Soo Joo, and Carlos Milla
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Agonist ,Carbachol ,Cystic Fibrosis ,Mucociliary clearance ,medicine.drug_class ,Swine ,Science ,Pharmacology ,Cystic fibrosis ,chemistry.chemical_compound ,medicine ,Animals ,Submucosal glands ,Multidisciplinary ,Forskolin ,Colforsin ,Ferrets ,Isoproterenol ,Drug Synergism ,Muscle, Smooth ,medicine.disease ,Disease Models, Animal ,chemistry ,Mucociliary Clearance ,Cholinergic ,Medicine ,Bronchoconstriction ,Drug Therapy, Combination ,medicine.symptom ,medicine.drug ,Muscle Contraction - Abstract
Mucus clearance, a primary innate defense mechanism of airways, is defective in patients with cystic fibrosis (CF) and CF animals. In previous work, the combination of a low dose of the cholinergic agonist, carbachol with forskolin or a β adrenergic agonist, isoproterenol synergistically increased mucociliary clearance velocity (MCCV) in ferret tracheas. Importantly, the present study shows that synergistic MCCV can also be produced in CF ferrets, with increases ~ 55% of WT. Synergistic MCCV was also produced in pigs. The combined agonists increased MCCV by increasing surface fluid via multiple mechanisms: increased fluid secretion from submucosal glands, increased anion secretion across surface epithelia and decreased Na+ absorption. To avoid bronchoconstriction, the cAMP agonist was applied 30 min before carbachol. This approach to increasing mucus clearance warrants testing for safety and efficacy in humans as a potential therapeutic for muco-obstructive diseases.
- Published
- 2021
4. Variation in CFTR-dependent ‘β-sweating’ among healthy adults
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Lesje DeRose, Jeeyeon Kim, Miesha Farahmand, Meagan Y. Shinbashi, Nam Soo Joo, and Jeffrey J. Wine
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Adult ,Multidisciplinary ,Cystic Fibrosis ,Cystic Fibrosis Transmembrane Conductance Regulator ,Humans ,Sweating ,Sweat ,Sweat Glands - Abstract
The genetic disease cystic fibrosis (CF) results when mutations in the gene for the anion channel CFTR reduce CFTR’s activity below a critical level. CFTR activity = N·PO·γ (number of channels x open probability x channel conductance). Small molecules are now available that partially restore CFTR function with dramatic improvements in health of CF subjects. Continued evaluation of these and other compounds in development will be aided by accurate assessments of CFTR function. However, measuring CFTR activity in vivo is challenging and estimates vary widely. The most accurate known measure of CFTR activity in vivo is the ‘β/M’ ratio of sweat rates, which is produced by stimulation with a β-adrenergic agonist cocktail referenced to the same individual’s methacholine-stimulated sweat rate. The most meaningful metric of CFTR activity is to express it as a percent of normal function, so it is critical to establish β/M carefully in a population of healthy control subjects. Here, we analyze β/M from a sample of 50 healthy adults in which sweat rates to cholinergic and β-adrenergic agonists were measured repeatedly (3 times) in multiple, (~50) identified sweat glands from each individual (giving ~20,000 measurements). The results show an approximately 7-fold range, 26–187% of the WT average set to 100%. These provide a benchmark against which other measures of CFTR activity can be compared. Factors contributing to β/M variation in healthy controls are discussed.
- Published
- 2022
5. 376: Synergistic mucociliary clearance in pig airways by beta-adrenergic and cholinergic agonists
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C. Milla, J. Wine, and Nam Soo Joo
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Pulmonary and Respiratory Medicine ,Adrenergic receptor ,Mucociliary clearance ,business.industry ,Pediatrics, Perinatology and Child Health ,medicine ,Cholinergic ,Pharmacology ,medicine.disease ,business ,Cystic fibrosis - Published
- 2021
6. Glandular Proteome Identifies Antiprotease Cystatin C as a Critical Modulator of Airway Hydration and Clearance
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Scott R. Tyler, Nam Soo Joo, Hyung Ju Cho, Jordy J. Hsiao, Michael E. Wright, T. Idil Apak Evans, Weiliang Xie, Yulong Zhang, Ziying Yan, Nicholas W. Keiser, John F. Engelhardt, and Jeffrey J. Wine
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0301 basic medicine ,Pulmonary and Respiratory Medicine ,Epithelial sodium channel ,medicine.medical_specialty ,Cystic Fibrosis ,Proteome ,Mucociliary clearance ,Clinical Biochemistry ,Bronchi ,Cystic fibrosis ,03 medical and health sciences ,Internal medicine ,medicine ,Animals ,Humans ,Cystatin C ,Molecular Biology ,Original Research ,Cathepsin S ,Submucosal glands ,biology ,Chemistry ,Ferrets ,Cell Biology ,respiratory system ,medicine.disease ,Cystic fibrosis transmembrane conductance regulator ,respiratory tract diseases ,Cell biology ,Trachea ,HEK293 Cells ,030104 developmental biology ,Endocrinology ,Chloride channel ,biology.protein - Abstract
Defects in the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel lead to viscous secretions from submucosal glands that cannot be properly hydrated and cleared by beating cilia in cystic fibrosis (CF) airways. The mechanisms by which CFTR, and the predominant epithelial sodium channel (ENaC), control the hydration and clearance of glandular secretions remain unclear. We used a proteomics approach to characterize the proteins contained in CF and non-CF submucosal gland fluid droplets and found that differentially regulated proteases (cathepsin S and H) and their antiprotease (cystatin C) influenced the equilibration of fluid on the airway surface and tracheal mucociliary clearance (MCC). Contrary to prevailing models of airway hydration and clearance, cystatin C, or raising the airway surface liquid (ASL) pH, inhibited cathepsin-dependent ENaC-mediated fluid absorption and raised the height of ASL, and yet decreased MCC velocity. Importantly, coupling of both CFTR and ENaC activities were required for effective MCC and for effective ASL height equilibration after volume challenge. Cystatin C–inhibitable cathepsins controlled initial phases of ENaC-mediated fluid absorption, whereas CFTR activity was required to prevent ASL dehydration. Interestingly, CF airway epithelia absorbed fluid more slowly owing to reduced cysteine protease activity in the ASL but became abnormally dehydrated with time. Our findings demonstrate that, after volume challenge, pH-dependent protease-mediated coupling of CFTR and ENaC activities are required for rapid fluid equilibration at the airway surface and for effective MCC. These findings provide new insights into how glandular fluid secretions may be equilibrated at the airway surface and how this process may be impaired in CF.
- Published
- 2016
7. Impaired PGE2-stimulated Cl- and HCO3- secretion contributes to cystic fibrosis airway disease
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Miriam F. Figueira, Zachary M. Sellers, Jeffrey J. Wine, Nam Soo Joo, Horst Fischer, Marcelo M. Morales, Jackson Souza-Menezes, Eric Sibley, Beate Illek, and Gopika Hari
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0301 basic medicine ,Cystic Fibrosis ,Pulmonology ,Physiology ,Respiratory System ,lcsh:Medicine ,Pathology and Laboratory Medicine ,Cystic fibrosis ,Epithelium ,Endocrinology ,Animal Cells ,Medicine and Health Sciences ,Prostaglandin E2 ,lcsh:Science ,Immune Response ,Cells, Cultured ,Submucosal glands ,Mammals ,Multidisciplinary ,Chemistry ,Niflumic acid ,food and beverages ,Eukaryota ,respiratory system ,Body Fluids ,Trachea ,Genetic Diseases ,Vertebrates ,Cellular Types ,Anatomy ,medicine.drug ,Research Article ,Mucociliary clearance ,Immunology ,Bronchi ,In Vitro Techniques ,Dinoprostone ,03 medical and health sciences ,Signs and Symptoms ,Autosomal Recessive Diseases ,Chlorides ,Diagnostic Medicine ,medicine ,Animals ,Humans ,Secretion ,Hormone transport ,Hormone Transport ,Clinical Genetics ,Inflammation ,Endocrine Physiology ,lcsh:R ,Organisms ,Ferrets ,Biology and Life Sciences ,Epithelial Cells ,Cell Biology ,medicine.disease ,Mucus ,Molecular biology ,Fibrosis ,Bicarbonates ,030104 developmental biology ,Biological Tissue ,Amniotes ,lcsh:Q ,Physiological Processes ,Developmental Biology - Abstract
Background Airway mucociliary clearance (MCC) is an important defense mechanism against pulmonary infections and is compromised in cystic fibrosis (CF). Cl- and HCO3- epithelial transport are integral to MCC. During pulmonary infections prostaglandin E2 (PGE2) production is abundant. Aim To determine the effect of PGE2 on airway Cl- and HCO3- secretion and MCC in normal and CF airways. Methods We examined PGE2 stimulated MCC, Cl- and HCO3- secretion using ferret trachea, human bronchial epithelial cell cultures (CFBE41o- with wildtype CFTR (CFBE41 WT) or homozygous F508del CFTR (CFBE41 CF) and human normal bronchial submucosal gland cell line (Calu-3) in Ussing chambers with or without pH-stat. Results PGE2 stimulated MCC in a dose-dependent manner and was partially impaired by CFTRinh-172. PGE2-stimulated Cl- current in ferret trachea was partially inhibited by CFTRinh-172, with niflumic acid eliminating the residual current. CFBE41 WT cell monolayers produced a robust Cl- and HCO3- secretory response to PGE2, both of which were completely inhibited by CFTRinh-172. CFBE41 CF cells exhibited no response to PGE2. In Calu-3 cells, PGE2 stimulated Cl- and HCO3- secretion. Cl- secretion was partially inhibited by CFTRinh-172, with additional inhibition by niflumic acid. HCO3- secretion was completely inhibited by CFTRinh-172. Conclusions PGE2 stimulates bronchotracheal MCC and this response is decreased in CF. In CF airway, PGE2-stimulated Cl- and HCO3- conductance is impaired and may contribute to decreased MCC. There remains a CFTR-independent Cl- current in submucosal glands, which if exploited, could represent a means of improving airway Cl- secretion and MCC in CF.
- Published
- 2017
8. Progress in understanding mucus abnormalities in cystic fibrosis airways
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Mario Pieper, Anna Ermund, Nam Soo Joo, Jeffrey J. Wine, Gunnar C. Hansson, and Peter König
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0301 basic medicine ,Pulmonary and Respiratory Medicine ,Pathology ,medicine.medical_specialty ,Cystic Fibrosis ,Mucociliary clearance ,Cystic Fibrosis Transmembrane Conductance Regulator ,Innate defence ,Cystic fibrosis ,Sterile environment ,03 medical and health sciences ,medicine ,Humans ,Submucosal glands ,business.industry ,respiratory system ,medicine.disease ,Mucus ,respiratory tract diseases ,Hypertonic saline ,030104 developmental biology ,Mucociliary Clearance ,Pediatrics, Perinatology and Child Health ,Immunology ,business ,Mucus clearance - Abstract
Normal airways below the carina maintain an essentially sterile environment via a multi-pronged innate defence system that includes mucus clearance via mucociliary clearance and cough, multiple antimicrobials and cellular components including macrophages and neutrophils. In cystic fibrosis (CF), loss of CFTR function compromises these defences, and with present standard of care virtually all people with CF eventually develop mucus accumulation, plugging and chronic infections. This review focuses on how mucus is affected by CFTR loss.
- Published
- 2017
9. Marked increases in mucociliary clearance produced by synergistic secretory agonists or inhibition of the epithelial sodium channel
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Hyung Ju Cho, Nam Soo Joo, Jin Hyeok Jeong, and Jeffrey J. Wine
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0301 basic medicine ,Epithelial sodium channel ,medicine.medical_specialty ,Carbachol ,Mucociliary clearance ,Cystic Fibrosis Transmembrane Conductance Regulator ,Article ,Amiloride ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Internal medicine ,Benzamil ,medicine ,Epithelial Sodium Channel Blockers ,Animals ,Epithelial Sodium Channels ,Multidisciplinary ,Forskolin ,Ion Transport ,biology ,Colforsin ,Ferrets ,Isoproterenol ,Drug Synergism ,respiratory system ,Cystic fibrosis transmembrane conductance regulator ,Up-Regulation ,030104 developmental biology ,Endocrinology ,chemistry ,Mucociliary Clearance ,biology.protein ,Chloride channel ,030217 neurology & neurosurgery ,medicine.drug - Abstract
Mucociliary clearance (MCC) is a critical host innate defense mechanism in airways, and it is impaired in cystic fibrosis (CF) and other obstructive lung diseases. Epithelial fluid secretion and absorption modify MCC velocity (MCCV). We tested the hypotheses that inhibiting fluid absorption accelerates MCCV, whereas inhibiting fluid secretion decelerates it. In airways, ENaC is mainly responsible for fluid absorption, while anion channels, including CFTR and Ca2+-activated chloride channels mediate anion/fluid secretion. MCCV was increased by the cAMP-elevating agonists, forskolin or isoproterenol (10 μM) and by the Ca2+-elevating agonist, carbachol (0.3 μM). The CFTR-selective inhibitor, CFTRinh-172, modestly reduced MCCV-increases induced by forskolin or isoproterenol but not increases induced by carbachol. The ENaC inhibitor benzamil increased basal MCCV as well as MCCV increases produced by forskolin or carbachol. MCC velocity was most dramatically accelerated by the synergistic combination of forskolin and carbachol, which produced near-maximal clearance rates regardless of prior treatment with CFTR or ENaC inhibitors. In CF airways, where CFTR-mediated secretion (and possibly synergistic MCC) is lost, ENaC inhibition via exogenous agents may provide therapeutic benefit, as has long been proposed.
- Published
- 2016
10. Properties of substance P-stimulated mucus secretion from porcine tracheal submucosal glands
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Jae Young Choi, Nam Soo Joo, Monal Khansaheb, Yu-Mi Yang, Jeffrey J. Wine, and Mauri E. Krouse
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Anions ,Pulmonary and Respiratory Medicine ,medicine.medical_specialty ,Pathology ,Time Factors ,Physiology ,Sus scrofa ,Cystic Fibrosis Transmembrane Conductance Regulator ,Neuropeptide ,Substance P ,Respiratory Mucosa ,In Vitro Techniques ,Biology ,Cystic fibrosis ,chemistry.chemical_compound ,Physiology (medical) ,Internal medicine ,medicine ,Animals ,Humans ,Microscopy, Interference ,Secretion ,Respiratory system ,Submucosal glands ,Cell Biology ,medicine.disease ,Mucus ,Trachea ,Endocrinology ,medicine.anatomical_structure ,chemistry ,Type C Phospholipases ,Calcium ,Carbachol ,Capsaicin ,Fura-2 ,Respiratory tract - Abstract
Human and pig airway submucosal glands secrete mucus in response to substance P (SubP), but in pig tracheal glands the response to SubP is >10-fold greater than in humans and shares features with cholinergically produced secretion. CFTR-deficient pigs provide a model for human cystic fibrosis (CF), and in newborn CF pigs the response of tracheal glands to SubP is significantly reduced (Joo et al. J Clin Invest 120: 3161–3166, 2010). To further define features of SubP-mediated gland secretion, we optically measured secretion rates from individual adult porcine glands in isolated tracheal tissues in response to mucosal capsaicin and serosal SubP. Mucosal capsaicin (EC50 = 19 μM) stimulated low rates of secretion that were partially inhibited by tetrodotoxin and by inhibitors for muscarinic, VIP, and SubP receptors, suggesting reflex stimulation of secretion by multiple transmitters. Secretion in response to mucosal capsaicin was inhibited by CFTRinh-172, but not by niflumic acid. Serosal SubP (EC50 = 230 nM) stimulated 10-fold more secretion than mucosal capsaicin, with a Vmax similar to that of carbachol. Secretion rates peaked within 5 min and then declined to a lower sustained rate. SubP-stimulated secretion was inhibited 75% by bumetanide, 53% by removal of HCO3−, and 85% by bumetanide + removal of HCO3−; it was not inhibited by atropine but was inhibited by niflumic acid, clotrimazole, BAPTA-AM, nominally Ca2+-free bath solution, and the adenylate cyclase inhibitor MDL-12330A. Ratiometric measurements of fura 2 fluorescence in dissociated gland cells showed that SubP and carbachol increased intracellular Ca2+ concentration by similar amounts. SubP produced rapid volume loss by serous and mucous cells, expansion of gland lumina, mucus flow, and exocytosis but little or no contraction of myoepithelial cells. These and prior results suggest that SubP stimulates pig gland secretion via CFTR- and Ca2+-activated Cl− channels.
- Published
- 2011
11. Hyposecretion of fluid from tracheal submucosal glands of CFTR-deficient pigs
- Author
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Monal Khansaheb, Jeffrey J. Wine, Nam Soo Joo, and Hyung Ju Cho
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Submucosal glands ,medicine.medical_specialty ,Pathology ,Cystic Fibrosis ,Swine ,Ferrets ,Cystic Fibrosis Transmembrane Conductance Regulator ,Biological Transport ,Mice, Transgenic ,General Medicine ,respiratory system ,Biology ,Epithelium ,respiratory tract diseases ,Animals, Genetically Modified ,Disease Models, Animal ,Mice ,Exocrine Glands ,Endocrinology ,Internal medicine ,medicine ,Animals ,Humans ,ΔF508 ,Research Article - Abstract
Cystic fibrosis (CF) results from mutations that disrupt CF transmembrane conductance regulator (CFTR), an anion channel found mainly in apical membranes of epithelial cells. CF leads to chronic infection of the airways with normally innocuous bacteria and fungi. Hypotheses to explain the pathophysiology of CF airways have been difficult to test because mouse models of CF do not develop human-like airway disease. The recent production of pigs lacking CFTR and pigs expressing the most common CF-causing CFTR mutant, ΔF508, provide another model that might help clarify the pathophysiology of CF airway disease. Here, we studied individual submucosal glands from 1-day-old piglets in situ in explanted tracheas, using optical methods to monitor mucus secretion rates from multiple glands in parallel. Secretion rates from control piglets (WT and CFTR+/–) and piglets with CF-like disease (CFTR–/– and CFTR–/ΔF508) were measured under 5 conditions: unstimulated (to determine basal secretion), stimulated with forskolin, stimulated with carbachol, stimulated with substance P, and, as a test for synergy, stimulated with forskolin and a low concentration of carbachol. Glands from piglets with CF-like disease responded qualitatively to all agonists like glands from human patients with CF, producing virtually no fluid in response to stimulation with forskolin and substantially less in response to all other agonists except carbachol. These data are a step toward determining whether gland secretory defects contribute to CF airway disease.
- Published
- 2010
12. Mucus secretion from individual submucosal glands of the ferret trachea
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Jeffrey J. Wine, Hyung Ju Cho, and Nam Soo Joo
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Anions ,Pulmonary and Respiratory Medicine ,Exocrine gland ,Pathology ,medicine.medical_specialty ,Swine ,Physiology ,Vasoactive intestinal peptide ,Cystic Fibrosis Transmembrane Conductance Regulator ,Cholinergic Agonists ,Phenylephrine ,Exocrine Glands ,Physiology (medical) ,medicine ,Animals ,Humans ,Vasoconstrictor Agents ,Secretion ,Respiratory system ,Submucosal glands ,Sheep ,biology ,Colforsin ,Ferrets ,Isoproterenol ,Drug Synergism ,Muscle, Smooth ,Articles ,Cell Biology ,Anatomy ,Mucus ,Cystic fibrosis transmembrane conductance regulator ,Bronchodilator Agents ,Trachea ,medicine.anatomical_structure ,biology.protein ,Carbachol ,Vasoactive Intestinal Peptide ,Respiratory tract - Abstract
Mucus secretion from individual tracheal glands in adult ferrets was studied with time-lapse optical imaging of mucus droplets under an oil layer. Density of functional glands (determined by responses to 1 μM carbachol) was 1.5 ± 0.3 per mm2( n = 6). Secretion rates (in pl·min−1·gland−1) were as follows: 4.1 ± 0.7 basal (unstimulated; n = 27, 669 glands), 338 ± 70 to 10 μM forskolin ( n = 8, 90 glands), 234 ± 13 to 1 μM VIP ( n = 6, 57 glands), 183 ± 92 to 10 μM isoproterenol ( n = 3, 33 glands), 978 ± 145 to 1 μM carbachol ( n = 11, 131 glands), and 1,348 ± 325 to 10 μM phenylephrine ( n = 7, 74 glands). The potency (EC50, in μM) and efficacy ( Vmax, in pl·min−1·gland−1) were 7.6 (EC50) and 338 ± 16 ( Vmax) to forskolin, 1.0 (EC50) and 479 ± 19 ( Vmax) to VIP, 0.6 (EC50) and 1,817 ± 268 ( Vmax) to carbachol, and 3.7 (EC50) and 1,801 ± 95 ( Vmax) to phenylephrine. Although carbachol and phenylephrine were equally effective secretagogues, only carbachol caused contractions of the trachealis muscle. Synergy was demonstrated between 300 nM isoproterenol and 100 nM carbachol, which, when combined, produced a secretion rate almost fourfold greater than predicted from their additive effect. The dependence of fluid secretion on Cl−and HCO3−varied depending on the mode of stimulation. Secretion stimulated by VIP or forskolin was reduced by ∼60% by blocking either anion, while carbachol-stimulated secretion was blocked 68% by bumetanide and only 32% by HEPES replacement of HCO3−. These results provide parametric data for comparison with fluid secretion from glands in ferrets lacking CFTR.
- Published
- 2010
13. Secretion rates of human nasal submucosal glands from patients with chronic rhinosinusitis or cystic fibrosis
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Nam Soo Joo, Jeffrey J. Wine, Peter H. Hwang, Do-Yeon Cho, and Jin Hyeok Jeong
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Adult ,Male ,Pathology ,medicine.medical_specialty ,Exocrine gland ,Cystic Fibrosis ,Mucous membrane of nose ,Cystic fibrosis ,Exocrine Glands ,medicine ,Immunology and Allergy ,Humans ,Nasal polyps ,Secretion ,Sinusitis ,Rhinitis ,Submucosal glands ,business.industry ,General Medicine ,Middle Aged ,medicine.disease ,Mucus ,Nasal Mucosa ,medicine.anatomical_structure ,Otorhinolaryngology ,Chronic Disease ,Female ,business - Abstract
Background A majority of patients with cystic fibrosis (CF) have chronic rhinosinusitis (CRS) and/or nasal polyps, both of which may be secondary to reduced fluid secretion from nasal submucosal glands. Objective To determine whether decreased fluid secretion from nasal submucosal glands also occurs in patients without CF and with CRS. Methods Inferior turbinates of the nasal cavity were harvested from controls, subjects with CRS, and subjects with CF (n = 5–7 per group). The secretion rates of the nasal submucosal glands of the three groups in response to carbachol and forskolin were measured by using time lapse digital imaging of mucus bubbles from single glands as they formed on the mucosal surface under oil. Results Carbachol-stimulated secretion rates were the following: controls, 1670 ± 381 pl-min−1-gland−1; CRS, 965 ± 440 pl-min−1-gland−1; and CF, 933 ± 588 pl-min−1-gland−1 (p = 0.23, Kruskal-Wallis test). Forskolin-stimulated secretion rates were the following: controls, 229 ± 14 pl-min−1-gland−1; CRS, 154 ± 48 pl-min−1-gland−1; and CF, 22 ± 15 pl-min−1-gland−1 (p = 0.008, Kruskal-Wallis test). The ratio of the average secretion rate induced by forskolin to that induced by carbachol was 13.7% in the controls, and 15.9% in CRS and 2.3% in CF groups. Conclusion The only significant difference in this small study was decreased forskolin-stimulated secretion in subjects with CF relative to the other subjects. However, there was a trend toward reduced carbachol-stimulated secretion rates in subjects with CRS and with and without CF relative to controls. Additional studies are needed to determine if nasal submucosal gland hyposecretion occurs in CRS either as a contributor to or as a consequence of CRS pathogenesis.
- Published
- 2015
14. Inhibition of airway surface fluid absorption by cholinergic stimulation
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Mauri E. Krouse, Jae Young Choi, Jeffrey J. Wine, Hyung Ju Cho, and Nam Soo Joo
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0301 basic medicine ,Epithelial sodium channel ,inorganic chemicals ,Atropine ,medicine.medical_specialty ,Carbachol ,Swine ,Stimulation ,Respiratory Mucosa ,Article ,Amiloride ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Internal medicine ,Benzamil ,medicine ,Animals ,Humans ,Secretion ,Epithelial Sodium Channels ,Cells, Cultured ,Multidisciplinary ,Sheep ,Ferrets ,respiratory system ,Mucus ,Trachea ,030104 developmental biology ,Endocrinology ,chemistry ,Cholinergic ,Rabbits ,030217 neurology & neurosurgery ,medicine.drug - Abstract
In upper airways airway surface liquid (ASL) depth and clearance rates are both increased by fluid secretion. Secretion is opposed by fluid absorption, mainly via the epithelial sodium channel, ENaC. In static systems, increased fluid depth activates ENaC and decreased depth inhibits it, suggesting that secretion indirectly activates ENaC to reduce ASL depth. We propose an alternate mechanism in which cholinergic input, which causes copious airway gland secretion, also inhibits ENaC-mediated absorption. The conjoint action accelerates clearance, and the increased transport of mucus out of the airways restores ASL depth while cleansing the airways. We were intrigued by early reports of cholinergic inhibition of absorption by airways in some species. To reinvestigate this phenomenon, we studied inward short-circuit currents (Isc) in tracheal mucosa from human, sheep, pig, ferret, and rabbit and in two types of cultured cells. Basal Isc was inhibited 20–70% by the ENaC inhibitor, benzamil. Long-lasting inhibition of ENaC-dependent Isc was also produced by basolateral carbachol in all preparations except rabbit and the H441 cell line. Atropine inhibition produced a slow recovery or prevented inhibition if added before carbachol. The mechanism for inhibition was not determined and is most likely multi-factorial. However, its physiological significance is expected to be increased mucus clearance rates in cholinergically stimulated airways.
- Published
- 2015
15. An Inwardly Rectifying Potassium Channel in Apical Membrane of Calu-3 Cells
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Jin V. Wu, Nam Soo Joo, Jeffrey J. Wine, Arjun Rustagi, and Mauri E. Krouse
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medicine.medical_specialty ,Patch-Clamp Techniques ,Potassium Channels ,Time Factors ,Cesium ,Cystic Fibrosis Transmembrane Conductance Regulator ,Biochemistry ,Cell Line ,Immediate-Early Proteins ,Adenosine Triphosphate ,Internal medicine ,medicine ,Humans ,Secretion ,RNA, Messenger ,Patch clamp ,Potassium Channels, Inwardly Rectifying ,Molecular Biology ,Monomeric GTP-Binding Proteins ,Dose-Response Relationship, Drug ,biology ,Reverse Transcriptase Polymerase Chain Reaction ,Cell Membrane ,Conductance ,Cell Biology ,Human airway ,Hydrogen-Ion Concentration ,Apical membrane ,Potassium channel ,Cystic fibrosis transmembrane conductance regulator ,Electrophysiology ,Kinetics ,Endocrinology ,Barium ,Permeability (electromagnetism) ,Potassium ,biology.protein ,Biophysics ,Chlorine - Abstract
Patch clamp methods and reverse transcription-polymerase chain reaction (RT-PCR) were used to characterize an apical K+ channel in Calu-3 cells, a widely used model of human airway gland serous cells. In cell-attached and excised apical membrane patches, we found an inwardly rectifying K+ channel (Kir). The permeability ratio was PNa/PK = 0.058. In 30 patches with both cystic fibrosis transmembrane conductance regulator and Kir present, we observed 79 cystic fibrosis transmembrane conductance regulator and 58 Kir channels. The average chord conductance was 24.4 +/- 0.5 pS (n = 11), between 0 and -200 mV, and was 9.6 +/- 0.7 pS (n = 8), between 0 and 50 mV; these magnitudes and their ratio of approximately 2.5 are most similar to values for rectifying K+ channels of the Kir4.x subfamilies. We attempted to amplify transcripts for Kir4.1, Kir4.2, and Kir5.1; of these only Kir4.2 was present in Calu-3 lysates. The channel was only weakly activated by ATP and was relatively insensitive to internal pH. External Cs+ and Ba2+ blocked the channel with Kd values in the millimolar range. Quantitative modeling of Cl- secreting epithelia suggests that secretion rates will be highest and luminal K+ will rise to 16-28 mm if 11-25% of the total cellular K+ conductance is placed in the apical membrane (Cook, D. I., and Young, J. A. (1989) J. Membr. Biol. 110, 139-146). Thus, we hypothesize that the K+ channel described here optimizes the rate of secretion and is involved in K+ recycling for the recently proposed apical H+ -K+ -ATPase in Calu-3 cells.
- Published
- 2004
16. Regulation of Antiprotease and Antimicrobial Protein Secretion by Airway Submucosal Gland Serous Cells
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Arjun Rustagi, Nam Soo Joo, Kimberly M. Winges, Dennis J. Lee, and Jeffrey J. Wine
- Subjects
Spectrometry, Mass, Electrospray Ionization ,medicine.medical_specialty ,Proteases ,Potassium Channels ,Time Factors ,Thapsigargin ,Swine ,alpha 1-Antichymotrypsin ,Molecular Sequence Data ,Anti-Inflammatory Agents ,Cystic Fibrosis Transmembrane Conductance Regulator ,Enzyme-Linked Immunosorbent Assay ,Respiratory Mucosa ,Biology ,Siderocalin ,Biochemistry ,Mass Spectrometry ,Cell Line ,chemistry.chemical_compound ,Internal medicine ,Glyburide ,medicine ,Animals ,Humans ,Protease Inhibitors ,Secretion ,Amino Acid Sequence ,Molecular Biology ,Chromatography, High Pressure Liquid ,Forskolin ,Sequence Homology, Amino Acid ,Reverse Transcriptase Polymerase Chain Reaction ,Colforsin ,Serine Endopeptidases ,Sodium ,Epithelial Cells ,Cell Biology ,Molecular biology ,Cystic fibrosis transmembrane conductance regulator ,Secretory protein ,Endocrinology ,chemistry ,alpha 1-Antitrypsin ,biology.protein ,Electrophoresis, Polyacrylamide Gel ,Muramidase ,Lysozyme ,Cell Division - Abstract
Airway submucosal gland serous cells express the cystic fibrosis transmembrane conductance regulator (CFTR) and secrete antimicrobial, anti-inflammatory, and antioxidant molecules. In cystic fibrosis, diminished gland secretion may impair innate airway host defenses. We used Calu-3 cells as a serous cell model to study the types of proteins released, the pathways that release them, and the possible involvement of CFTR activity in protein release. Many proteins were secreted constitutively into the apical fluid and showed increased release to agonists. We identified some of them by high pressure liquid chromatography-mass spectrometry and reverse transcriptase PCR, including lysozyme, siderocalin (the protein NGAL), which inhibits bacterial growth by binding iron-containing siderophores, HSC-71, which is thought to have anti-inflammatory properties, and the serine protease inhibitors alpha-1-antitrypsin and alpha-1-antichymotrypsin, which may function as antimicrobials as well as play a potential role in diminishing the activation of epithelial Na(+) channels by serine proteases. We used an enzyme-linked immunosorbent assay to quantify lysozyme secretion by Calu-3 cells in response to various agonists and inhibitors. Forskolin increased the lysozyme secretion rate (J(lyz)) from 32 to 77 ng/hr/cm(2) (n = 36, p < 0.005). Thapsigargin increased J(lyz) from 40 to 63 ng/h/cm(2) (n = 16, p < 0.005), and forskolin plus thapsigargin further increased the forskolin-stimulated J(lyz) by 48% (n = 9, p < 0.05). 1-Ethyl-benzimidazolinone and carbachol were less effective. Glibenclamide inhibited basal and stimulated J(lyz), but clotrimazole was without effect. CFTR(inh)172 caused a small (15%) but significant inhibition of forskolin-stimulated J(lyz) without affecting basal J(lyz). Thus, Calu-3 cells secrete diverse proteins that in aggregate would be expected to suppress microbial growth, protect the airways from damage, and limit the activation of epithelial Na(+) channels via serine proteases.
- Published
- 2004
17. Numerical Study of Flow Around Impulsively Started Elliptic Cylinder Using Vortex Particle Method
- Author
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Sang-Hwan Lee and Nam-Soo Joo
- Subjects
Convection ,Physics ,Field (physics) ,Mechanical Engineering ,Reynolds number ,Mechanics ,Vortex ,Physics::Fluid Dynamics ,symbols.namesake ,Classical mechanics ,Flow (mathematics) ,Vorticity equation ,symbols ,Burgers vortex ,Boundary value problem - Abstract
In this paper, the unsteady behavior of the viscous flow field past an impulsively started elliptic cylinder is studied numerically. In order to analyze flow field, we introduce vortex particle method. The vorticity transport equation is solved by fractional step algorithm which splits into convection term and diffusion term. The convection term is calculated with Biot-Savart law, the no-through boundary condition is employed on solid boundaries. The diffusion term is modified based on the scheme of particle strength exchange. The particle redistributed scheme for general geometry is adapted. The flows around an elliptic cylinder are investigated for various attack angles at Reynolds number 200. The comparison between numerical results of present study and experimental data shows good agreements.
- Published
- 2004
18. Absent Secretion to Vasoactive Intestinal Peptide in Cystic Fibrosis Airway Glands
- Author
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Robert C. Robbins, Nam Soo Joo, Richard I. Whyte, Jeffrey J. Wine, Jin V. Wu, and Toshiya Irokawa
- Subjects
Adult ,Male ,medicine.medical_specialty ,Cystic Fibrosis ,Biopsy ,Vasoactive intestinal peptide ,Bronchi ,Respiratory Mucosa ,Biology ,Biochemistry ,Cystic fibrosis ,stomatognathic system ,Internal medicine ,medicine ,Humans ,Secretion ,Molecular Biology ,Submucosal glands ,Lung ,Colforsin ,Myoepithelial cell ,Cell Biology ,Middle Aged ,respiratory system ,medicine.disease ,Mucus ,Tissue Donors ,Trachea ,Kinetics ,Serous fluid ,Endocrinology ,medicine.anatomical_structure ,Carbachol ,Female ,Lung Transplantation ,Vasoactive Intestinal Peptide - Abstract
We are testing the hypothesis that the malfunctioning of airway gland serous cells is a component of cystic fibrosis (CF) airway disease. CF is caused by mutations that disrupt CF transmembrane conductance regulator, an anion channel essential for proper fluid secretion in some epithelia. Submucosal glands supply most of the mucus in upper airways, and gland serous cells are the primary site of CF transmembrane conductance regulator expression in airways. We have discovered a major defect in CF glands by in situ optical monitoring of secretions from single human airway glands. CF glands did not secrete to agents that elevated [cAMP](i) (0 responses/450 glands, 8 subjects), whereas glands were responsive in all donor tracheas (605/827 glands, 15 subjects) and in bronchi from subjects who were transplanted because of other lung diseases (148/166 glands, n = 10). CF glands secreted to cholinergic stimulation, and serous cells were abundant in glands from all CF subjects. The complete absence of secretion to agents that elevate [cAMP](i) suggests that altered secretion of gland mucus could contribute to CF lung disease.
- Published
- 2002
19. Mucus Secretion from Single Submucosal Glands of Pig
- Author
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Yamil Saenz, Jeffrey J. Wine, Nam Soo Joo, and Mauri E. Krouse
- Subjects
Submucosal glands ,medicine.medical_specialty ,Thapsigargin ,Carbachol ,Forskolin ,Vasoactive intestinal peptide ,Cell Biology ,Biology ,Biochemistry ,Mucus ,chemistry.chemical_compound ,Endocrinology ,chemistry ,Internal medicine ,medicine ,Secretion ,Molecular Biology ,Bumetanide ,medicine.drug - Abstract
Secretion rates of >700 individual glands in isolated tracheal mucosa from 56 adult pigs were monitored optically. "Basal" secretion of 0.7 +/- 0.1 nl x min(-1) gland(-1) was observed 1-9 h post-harvest but was near zero on day 2. Secretion to carbachol (10 microm) peaked at 2-3 min and then declined to a sustained phase. Peak secretion was 12.4 +/- 1.1 nl x min(-1) gland(-1); sustained secretion was approximately one-third of peak secretion. Thapsigargin (1 microm) increased secretion from 0.1 +/- 0.05 to 0.7 +/- 0.2 nl x min(-1) gland(-1); thapsigargin did not cause contraction of the trachealis muscles. Isoproterenol and phenylephrine (10 microm each) were ineffective, but vasoactive intestinal peptide (1 microm) and forskolin (10 microm) each produced sustained secretion of 1.0 +/- 0.5 and 1.7 +/- 0.2 nl x min(-1) gland(-1), respectively. The density of actively secreting glands was 1.3/mm(2). Secretion to either carbachol or forskolin was inhibited (approximately 50%) by either bumetanide or HCO(3)(-) removal and inhibited approximately 90% by the combined treatments. Mucus secreted in response to carbachol or forskolin was acidic by approximately 0.2 pH units relative to the bath and remained acidic by approximately 0.1 pH units after bumetanide. The strong secretory response to vasoactive intestinal peptide, the acidity of [cAMP](i)-stimulated mucus, and its inhibition by bumetanide were unexpected.
- Published
- 2002
20. Mucociliary clearance and submucosal gland secretion in the ex vivo ferret trachea
- Author
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Peter H. Hwang, Jin Hyeok Jeong, Nam Soo Joo, and Jeffrey J. Wine
- Subjects
Pulmonary and Respiratory Medicine ,medicine.medical_specialty ,Carbachol ,Cystic Fibrosis ,Physiology ,Mucociliary clearance ,Vasoactive intestinal peptide ,Carbonates ,Cystic Fibrosis Transmembrane Conductance Regulator ,Angiogenesis Inhibitors ,chemistry.chemical_compound ,Exocrine Glands ,Chlorides ,Sodium Potassium Chloride Symporter Inhibitors ,Physiology (medical) ,Internal medicine ,medicine ,Cyclic AMP ,Animals ,Phenylephrine ,Bumetanide ,Submucosal glands ,Forskolin ,Ion Transport ,Colforsin ,Ferrets ,Isoproterenol ,food and beverages ,Cell Biology ,Analgesics, Non-Narcotic ,Mucus ,Bronchodilator Agents ,Trachea ,Endocrinology ,chemistry ,Mucociliary Clearance ,Nitrobenzoates ,Calcium ,Female ,medicine.drug - Abstract
In many species submucosal glands are an important source of tracheal mucus, but the extent to which mucociliary clearance (MCC) depends on gland secretion is unknown. To explore this relationship, we measured basal and agonist-stimulated MCC velocities in ex vivo tracheas from adult ferrets and compared the velocities with previously measured rates of ferret glandular mucus secretion (Cho HJ, Joo NS, Wine JJ. Am J Physiol Lung Cell Mol Physiol 299: L124–L136, 2010). Stimulated MCC velocities (mm/min, means ± SE for 10- to 35-min period poststimulation) were as follows: 1 μM carbachol: 19.1 ± 3.3 > 10 μM phenylephrine: 15.3 ± 2.4 ≈ 10 μM isoproterenol: 15.0 ± 1.9 ≈ 10 μM forskolin: 14.6 ± 3.1 > 1 μM vasoactive intestinal peptide (VIP): 10.2 ± 2.2 >> basal ( t15): 1.8 ± 0.3; n = 5–10 for each condition. Synergistic stimulation of MCC was observed between low concentrations of carbachol (100 nM) and isoproterenol (300 nM). Bumetanide inhibited carbachol-stimulated MCC by ∼70% and abolished the increase in MCC stimulated by forskolin + VIP, whereas HCO3−-free solutions did not significantly inhibit MCC to either intracellular Ca2+concentration or intracellular cAMP concentration ([cAMP]i)-elevating agonists. Stimulation and inhibition of MCC and gland secretion differed in several respects: most importantly, elevating [cAMP]iincreased MCC much more effectively than expected from its effects on gland secretion, and bumetanide almost completely inhibited [cAMP]i-stimulated MCC while it had a smaller effect on gland secretion. We conclude that changes in glandular fluid secretion are complexly related to MCC and discuss possible reasons for this.
- Published
- 2014
21. Optical method for quantifying rates of mucus secretion from single submucosal glands
- Author
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Mauri E. Krouse, Nam Soo Joo, Yamil Saenz, Jin V. Wu, and Jeffrey J. Wine
- Subjects
Male ,Pulmonary and Respiratory Medicine ,Optics and Photonics ,Exocrine gland ,Swine ,Physiology ,Mucociliary clearance ,Cholinergic Agonists ,Biology ,Tracheal mucosa ,stomatognathic system ,Physiology (medical) ,Submucosa ,Methods ,medicine ,Animals ,Secretion ,Bumetanide ,Submucosal glands ,Mucous Membrane ,Drug Synergism ,Cell Biology ,Anatomy ,Mucus ,Trachea ,Bicarbonates ,medicine.anatomical_structure ,Cats ,Biophysics ,Carbachol ,Female ,Respiratory tract - Abstract
We describe an optical method to quantify single- gland secretion. Isolated tracheal mucosa were mounted at the air-Krebs interface and coated with oil. Gland secretions formed spherical bubbles that were digitally imaged at intervals, allowing rates of secretion to be calculated. We monitored 340 glands in 54 experiments with 12 sheep. Glands secreted basally at low rates (0.57 ± 0.04 nl · min−1 · gland−1, 123 glands) in tissues up to 9 h postharvest and at lower rates for up to 3 days. Carbachol (10 μM) stimulated secretion with an early transient and a sustained or oscillating phase. Peak secretion was 15.7 ± 1.2 nl · min−1 · gland−1 (60 glands); sustained secretion was 4.5 ± 0.5 nl · min−1 · gland−1 (10 glands). Isoproterenol and phenylephrine (10 μM each) stimulated only small, transient responses. We confirmed that cats have a large secretory response to phenylephrine (11.6 ± 3.7 nl · min−1 · gland−1, 12 glands), but pigs, sheep, and humans all have small responses (−1 · gland−1). Carbachol-stimulated peak secretion was inhibited 56% by bumetanide, 67% by HCO[Formula: see text] replacement with HEPES, and 92% by both. The distribution of secretion rates was nonnormal, suggesting the existence of subpopulations of glands.
- Published
- 2001
22. Cystic Fibrosis Transmembrane Conductance Regulator Gating Requires Cytosolic Electrolytes
- Author
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Jeffrey J. Wine, Mauri E. Krouse, Jin V. Wu, and Nam Soo Joo
- Subjects
congenital, hereditary, and neonatal diseases and abnormalities ,Cystic Fibrosis Transmembrane Conductance Regulator ,ATP-binding cassette transporter ,Gating ,Biochemistry ,Cystic fibrosis ,Electrolytes ,Adenosine Triphosphate ,Cytosol ,medicine ,Humans ,Phosphorylation ,Molecular Biology ,Cells, Cultured ,biology ,Chemistry ,Cell Biology ,Hydrogen-Ion Concentration ,medicine.disease ,Adenosine ,Cystic fibrosis transmembrane conductance regulator ,Osmolyte ,Biophysics ,biology.protein ,Ion Channel Gating ,medicine.drug - Abstract
Cystic fibrosis transmembrane conductance regulator (CFTR), which causes cystic fibrosis when nonfunctional, is an anion channel and a member of the ATP binding cassette superfamily. After phosphorylation, CFTR gates by binding and hydrolyzing ATP. We show that CFTR open probability (P(o)) also depends on the electrolyte concentration of the cytosol. Inside-out patches from Calu-3 cells were transiently exposed to solutions of 160 mm salt or solutions in which up to 90% of the salt was replaced by nonionic osmolytes such as sucrose. In lowered salt solutions, CFTR P(o) declined within 1 s to a stable lower value that depended on the electrolyte concentration, (K(1/2) approximately 80 mm NaCl). P(o) was rapidly restored in normal salt concentrations without regard to the electrolyte species. Reducing external electrolytes did not affect CFTR P(o). The same results were obtained when CFTR was stably phosphorylated with adenosine 5'-O-(thiotriphosphate). The decrease in P(o) resulted entirely from an increase in mean closed time. Increasing ATP levels up to 20-fold did not counteract the effect of low electrolytes. The same effect was observed for CFTR expressed in C127 cells but not for a different species of anion channel. Cytosolic electrolytes are an unsuspected, essential cofactor for CFTR gating.
- Published
- 2001
23. Cloning of ClC-2 chloride channel from murine duodenum and its presence in CFTR knockout mice
- Author
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Byung Hee Han, Hyun Dju Kim, Lane L. Clarke, Nam Soo Joo, and Leonard R. Forte
- Subjects
DNA, Complementary ,Duodenum ,Molecular Sequence Data ,Biophysics ,Cystic Fibrosis Transmembrane Conductance Regulator ,Biochemistry ,Mice ,Rapid amplification of cDNA ends ,Intestinal mucosa ,Chloride Channels ,Structural Biology ,Complementary DNA ,Genetics ,Animals ,Amino Acid Sequence ,Cloning, Molecular ,Intestinal Mucosa ,Peptide sequence ,Mice, Knockout ,Cloning ,chemistry.chemical_classification ,Base Sequence ,biology ,Reverse Transcriptase Polymerase Chain Reaction ,urogenital system ,Blotting, Northern ,Molecular biology ,Cystic fibrosis transmembrane conductance regulator ,Amino acid ,CLC-2 Chloride Channels ,chemistry ,Chloride channel ,biology.protein ,RNA ,Sequence Alignment ,Plasmids - Abstract
We report the cloning of a murine ClC-2 chloride channel cDNA from duodenal epithelium by reverse transcriptase-polymerase chain reaction (RT-PCR) using degenerate primers and by rapid amplification of cDNA ends (RACE)-PCR. Other than CFTR, this represents the first cloned chloride channel from intact intestine. The ClC-2 cDNA predicts encoding of a 908 amino acid polypeptide with a calculated M(r) of 99,373. The amino acid sequence of the murine ClC-2 chloride channel is over 94% identical to the ClC-2 chloride channel proteins of other species. Of interest is the finding that the ClC-2 mRNA is expressed about the same level in duodena from both CFTR knockout and wild-type mice. This is in keeping with the suggestion that ClC-2 might be a therapeutic target in cystic fibrosis.
- Published
- 1999
24. Defective Fluid Secretion from Submucosal Glands of Nasal Turbinates from CFTR-/- and CFTRΔF508/ΔF508 Pigs
- Author
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Nam Soo Joo, Jeffrey J. Wine, and Hyung Ju Cho
- Subjects
Exocrine gland ,Aging ,Anatomy and Physiology ,Cystic Fibrosis ,Pulmonology ,Respiratory System ,Sus scrofa ,lcsh:Medicine ,Cystic Fibrosis Transmembrane Conductance Regulator ,Mucous membrane of nose ,Substance P ,Turbinates ,chemistry.chemical_compound ,0302 clinical medicine ,Autosomal Recessive ,Comparative Anatomy ,lcsh:Science ,Submucosal glands ,0303 health sciences ,Multidisciplinary ,Forskolin ,Drug Synergism ,Nasal glands ,Animal Models ,Organ Size ,respiratory system ,3. Good health ,Body Fluids ,Trachea ,medicine.anatomical_structure ,Medicine ,Research Article ,medicine.medical_specialty ,Biology ,03 medical and health sciences ,Model Organisms ,Exocrine Glands ,Internal medicine ,medicine ,Genetics ,Upper Respiratory Tract Infections ,Animals ,030304 developmental biology ,Clinical Genetics ,lcsh:R ,Colforsin ,Human Genetics ,Rhinology ,Mucus ,Endocrinology ,chemistry ,Otorhinolaryngology ,Animals, Newborn ,Respiratory Infections ,Mutation ,Nasal Diseases ,lcsh:Q ,Carbachol ,Fluid Physiology ,Nasal concha ,030217 neurology & neurosurgery - Abstract
Background Cystic fibrosis (CF), caused by reduced CFTR function, includes severe sinonasal disease which may predispose to lung disease. Newly developed CF pigs provide models to study the onset of CF pathophysiology. We asked if glands from pig nasal turbinates have secretory responses similar to those of tracheal glands and if CF nasal glands show reduced fluid secretion. Methodology/Principal Findings Unexpectedly, we found that nasal glands differed from tracheal glands in five ways, being smaller, more numerous (density per airway surface area), more sensitive to carbachol, more sensitive to forskolin, and nonresponsive to Substance P (a potent agonist for pig tracheal glands). Nasal gland fluid secretion from newborn piglets (12 CF and 12 controls) in response to agonists was measured using digital imaging of mucus bubbles formed under oil. Secretion rates were significantly reduced in all conditions tested. Fluid secretory rates (Controls vs. CF, in pl/min/gland) were as follows: 3 µM forskolin: 9.2±2.2 vs. 0.6±0.3; 1 µM carbachol: 143.5±35.5 vs. 52.2±10.3; 3 µM forskolin + 0.1 µM carbachol: 25.8±5.8 vs. CF 4.5±0.9. We also compared CFΔF508/ΔF508 with CFTR-/- piglets and found significantly greater forskolin-stimulated secretion rates in the ΔF508 vs. the null piglets (1.4±0.8, n = 4 vs. 0.2±0.1, n = 7). An unexpected age effect was also discovered: the ratio of secretion to 3 µM forskolin vs. 1 µM carbachol was ∼4 times greater in adult than in neonatal nasal glands. Conclusions/Significance These findings reveal differences between nasal and tracheal glands, show defective fluid secretion in nasal glands of CF pigs, reveal some spared function in the ΔF508 vs. null piglets, and show unexpected age-dependent differences. Reduced nasal gland fluid secretion may predispose to sinonasal and lung infections.
- Published
- 2011
25. Measurement of fluid secretion from intact airway submucosal glands
- Author
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Jeffrey J, Wine, Nam Soo, Joo, Jae Young, Choi, Hyung-Ju, Cho, Mauri E, Krouse, Jin V, Wu, Monal, Khansaheb, Toshiya, Irokawa, Juan, Ianowski, John W, Hanrahan, Alan W, Cuthbert, and Kim V, Tran
- Subjects
Microscopy ,Mucous Membrane ,Sheep ,Cystic Fibrosis ,Swine ,Ferrets ,Sputum ,Cystic Fibrosis Transmembrane Conductance Regulator ,Biological Transport ,Respiratory Mucosa ,Body Fluids ,Molecular Imaging ,Trachea ,Mice ,Mucus ,Exocrine Glands ,Species Specificity ,Mucociliary Clearance ,Cats ,Animals ,Humans - Abstract
Human airways are kept sterile by a mucosal innate defense system that includes mucus secretion. Mucus is secreted in healthy upper airways primarily by submucosal glands and consists of defense molecules mixed with mucins, electrolytes, and water and is also a major component of sputum. Mucus traps pathogens and mechanically removes them via mucociliary clearance while inhibiting their growth via molecular (e.g., lysozyme) and cellular (e.g., neutrophils, macrophages) defenses. Fluid secretion rates of single glands in response to various mediators can be measured by trapping the primary gland mucus secretions in an oil layer, where they form spherical bubbles that can be optically measured at any desired interval to provide detailed temporal analysis of secretion rates. The composition and properties of the mucus (e.g., solids, viscosity, pH) can also be determined. These methods have now been applied to mice, ferrets, cats, pigs, sheep, and humans, with a main goal of comparing gland secretion in control and CFTR-deficient humans and animals.
- Published
- 2011
26. Measurement of Fluid Secretion from Intact Airway Submucosal Glands
- Author
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Mauri E. Krouse, Hyung Ju Cho, Nam Soo Joo, Alan W. Cuthbert, Kim V. Tran, Jeffrey J. Wine, Toshiya Irokawa, Juan P. Ianowski, John W. Hanrahan, Jin V. Wu, Monal Khansaheb, and Jae Young Choi
- Subjects
Submucosal glands ,CATS ,Chemistry ,Mucociliary clearance ,Mucin ,respiratory system ,Mucus ,Cell biology ,chemistry.chemical_compound ,medicine ,Sputum ,Secretion ,Lysozyme ,medicine.symptom - Abstract
Human airways are kept sterile by a mucosal innate defense system that includes mucus secretion. Mucus is secreted in healthy upper airways primarily by submucosal glands and consists of defense molecules mixed with mucins, electrolytes, and water and is also a major component of sputum. Mucus traps pathogens and mechanically removes them via mucociliary clearance while inhibiting their growth via molecular (e.g., lysozyme) and cellular (e.g., neutrophils, macrophages) defenses. Fluid secretion rates of single glands in response to various mediators can be measured by trapping the primary gland mucus secretions in an oil layer, where they form spherical bubbles that can be optically measured at any desired interval to provide detailed temporal analysis of secretion rates. The composition and properties of the mucus (e.g., solids, viscosity, pH) can also be determined. These methods have now been applied to mice, ferrets, cats, pigs, sheep, and humans, with a main goal of comparing gland secretion in control and CFTR-deficient humans and animals.
- Published
- 2011
27. The mechanism of PAR-2 mediated mucus secretion in airway submucosal gland
- Author
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Nam Soo Joo, D.-M. Shin, H.-J. Cho, J.-Y. Choi, and H.-J. Lee
- Subjects
Pulmonary and Respiratory Medicine ,Mechanism (biology) ,business.industry ,respiratory system ,medicine.disease ,Cystic fibrosis ,Mucus ,Cell biology ,respiratory tract diseases ,Pediatrics, Perinatology and Child Health ,medicine ,Secretion ,Pediatrics, Perinatology, and Child Health ,business ,Airway - Published
- 2010
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28. Disease phenotype of a ferret CFTR-knockout model of cystic fibrosis
- Author
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Hongshu Sui, Jeffrey J. Wine, Paul W. Naumann, Weihong Zhou, Xingshen Sun, Yaling Yi, Timothy S. Frana, Xiaoming Liu, Michelle Griffin, Yulong Zhang, Joann M. Kinyon, Ziying Yan, Diana C.M. Lei-Butters, Hyung Ju Cho, Nam Soo Joo, John T. Fisher, John F. Engelhardt, Kai Wang, David K. Meyerholz, Jill Ascher, and Meihui Luo
- Subjects
Male ,congenital, hereditary, and neonatal diseases and abnormalities ,Pathology ,medicine.medical_specialty ,Malabsorption ,Cystic Fibrosis ,Respiratory System ,Clone (cell biology) ,Meconium Ileus ,Cystic Fibrosis Transmembrane Conductance Regulator ,Disease ,Biology ,Cystic fibrosis ,Vas Deferens ,medicine ,Animals ,Humans ,Lung ,Cells, Cultured ,Ferrets ,General Medicine ,respiratory system ,medicine.disease ,Phenotype ,Ursodeoxycholic acid ,respiratory tract diseases ,Disease Models, Animal ,medicine.anatomical_structure ,Technical Advance ,Animals, Newborn ,Immunology ,Mutation ,Commentary ,medicine.drug - Abstract
Cystic fibrosis (CF) is a recessive disease that affects multiple organs. It is caused by mutations in CFTR. Animal modeling of this disease has been challenging, with species- and strain-specific differences in organ biology and CFTR function influencing the emergence of disease pathology. Here, we report the phenotype of a CFTR-knockout ferret model of CF. Neonatal CFTR-knockout ferrets demonstrated many of the characteristics of human CF disease, including defective airway chloride transport and submucosal gland fluid secretion; variably penetrant meconium ileus (MI); pancreatic, liver, and vas deferens disease; and a predisposition to lung infection in the early postnatal period. Severe malabsorption by the gastrointestinal (GI) tract was the primary cause of death in CFTR-knockout kits that escaped MI. Elevated liver function tests in CFTR-knockout kits were corrected by oral administration of ursodeoxycholic acid, and the addition of an oral proton-pump inhibitor improved weight gain and survival. To overcome the limitations imposed by the severe intestinal phenotype, we cloned 4 gut-corrected transgenic CFTR-knockout kits that expressed ferret CFTR specifically in the intestine. One clone passed feces normally and demonstrated no detectable ferret CFTR expression in the lung or liver. The animals described in this study are likely to be useful tools for dissecting CF disease pathogenesis and developing treatments.
- Published
- 2010
29. Substance P stimulates human airway submucosal gland secretion mainly via a CFTR-dependent process
- Author
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Jeffrey J. Wine, Nam Soo Joo, Jae Young Choi, Monal Khansaheb, David Weill, Robert C. Robbins, and Mauri E. Krouse
- Subjects
Male ,medicine.medical_specialty ,Exocrine gland ,Cystic Fibrosis ,Vasoactive intestinal peptide ,Sus scrofa ,Cystic Fibrosis Transmembrane Conductance Regulator ,Substance P ,In Vitro Techniques ,Cystic fibrosis ,chemistry.chemical_compound ,Exocrine Glands ,stomatognathic system ,Internal medicine ,medicine ,Animals ,Humans ,Plant Oils ,Secretion ,Calcium Signaling ,Clotrimazole ,Egtazic Acid ,Chelating Agents ,Submucosal glands ,biology ,Dose-Response Relationship, Drug ,business.industry ,Colforsin ,Age Factors ,Drug Synergism ,General Medicine ,respiratory system ,medicine.disease ,Mucus ,Cystic fibrosis transmembrane conductance regulator ,Trachea ,Endocrinology ,medicine.anatomical_structure ,chemistry ,biology.protein ,Carbachol ,Female ,business ,Corrigendum ,Capsicum ,Vasoactive Intestinal Peptide ,Research Article - Abstract
Chronic bacterial airway infections are the major cause of mortality in cystic fibrosis (CF). Normal airway defenses include reflex stimulation of submucosal gland mucus secretion by sensory neurons that release substance P (SubP). CFTR is an anion channel involved in fluid secretion and mutated in CF; the role of CFTR in secretions stimulated by SubP is unknown. We used optical methods to measure SubP-mediated secretion from human submucosal glands in lung transplant tissue. Glands from control but not CF subjects responded to mucosal chili oil. Similarly, serosal SubP stimulated secretion in more than 60% of control glands but only 4% of CF glands. Secretion triggered by SubP was synergistic with vasoactive intestinal peptide and/or forskolin but not with carbachol; synergy was absent in CF glands. Pig glands demonstrated a nearly 10-fold greater response to SubP. In 10 of 11 control glands isolated by fine dissection, SubP caused cell volume loss, lumen expansion, and mucus flow, but in 3 of 4 CF glands, it induced lumen narrowing. Thus, in CF, the reduced ability of mucosal irritants to stimulate airway gland secretion via SubP may be another factor that predisposes the airways to infections.
- Published
- 2008
30. Synergistic airway gland mucus secretion in response to vasoactive intestinal peptide and carbachol is lost in cystic fibrosis
- Author
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Nam Soo Joo, Mauri E. Krouse, Jeffrey J. Wine, Jae Young Choi, Jin V. Wu, John W. Hanrahan, Robert C. Robbins, and Juan P. Ianowski
- Subjects
Exocrine gland ,medicine.medical_specialty ,Carbachol ,Cystic Fibrosis ,Swine ,Vasoactive intestinal peptide ,Respiratory System ,Cystic Fibrosis Transmembrane Conductance Regulator ,Biology ,Cholinergic Agonists ,Cystic fibrosis ,Exocrine Glands ,stomatognathic system ,Internal medicine ,medicine ,Cyclic AMP ,Animals ,Humans ,Secretion ,Drug Synergism ,General Medicine ,respiratory system ,medicine.disease ,Mucus ,Cystic fibrosis transmembrane conductance regulator ,Acetylcholine ,medicine.anatomical_structure ,Endocrinology ,biology.protein ,Cholinergic ,medicine.drug ,Vasoactive Intestinal Peptide ,Research Article - Abstract
Cystic fibrosis (CF) is caused by dysfunction of the CF transmembrane conductance regulator (CFTR), an anion channel whose dysfunction leads to chronic bacterial and fungal airway infections via a pathophysiological cascade that is incompletely understood. Airway glands, which produce most airway mucus, do so in response to both acetylcholine (ACh) and vasoactive intestinal peptide (VIP). CF glands fail to secrete mucus in response to VIP, but do so in response to ACh. Because vagal cholinergic pathways still elicit strong gland mucus secretion in CF subjects, it is unclear whether VIP-stimulated, CFTR-dependent gland secretion participates in innate defense. It was recently hypothesized that airway intrinsic neurons, which express abundant VIP and ACh, are normally active and stimulate low-level gland mucus secretion that is a component of innate mucosal defenses. Here we show that low levels of VIP and ACh produced significant mucus secretion in human glands via strong synergistic interactions; synergy was lost in glands of CF patients. VIP/ACh synergy also existed in pig glands, where it was CFTR dependent, mediated by both Cl(-) and HCO(3) (-), and clotrimazole sensitive. Loss of "housekeeping" gland mucus secretion in CF, in combination with demonstrated defects in surface epithelia, may play a role in the vulnerability of CF airways to bacterial infections.
- Published
- 2007
31. 39 Inhibition of CFTR slows forskolin-stimulated mucociliary clearance in ferret trachea
- Author
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Nam Soo Joo, Jeffrey J. Wine, and J.H. Jeong
- Subjects
Pulmonary and Respiratory Medicine ,Agonist ,medicine.medical_specialty ,Carbachol ,Forskolin ,Mucociliary clearance ,business.industry ,medicine.drug_class ,food and beverages ,respiratory system ,medicine.disease ,Mucus ,Cystic fibrosis ,chemistry.chemical_compound ,Endocrinology ,chemistry ,Internal medicine ,Pediatrics, Perinatology and Child Health ,medicine ,Secretion ,business ,Ex vivo ,medicine.drug - Abstract
Objective Mucus clearance is a primary defense mechanism of the airways. Mucus secretion and clearance are abnormal in diverse organs in humans with CF and in all CF animal models. To better understand the contribution of CFTR to mucociliary clearance (MCC), we asked if CFTR plays a role in ferret tracheal MCC by directly inhibiting CFTR with a selective CFTR inhibitor, CFTRinh172. Methods We measured MCC velocities in ex vivo adult ferret tracheas by following particle movements with time-lapse digital imaging (Jeong et al . 2014 AJP-lung, L83) in the presence or absence of CFTRinh172. Tracheas were mounted side-by side in a chamber that was sealed, 37°C-maintained, and continuously supplied with warmed, humidified 95% O 2 -5% CO 2 gas. Particle movements (MCC) were measured at 20 sec intervals for 30 min: 10 mM forskolin or 0.3 mM carbachol +/– CFTRinh172. Tissue viability was assessed by then stimulating MCC for 30 min with 0.3 mM carbachol plus 10 mM forskolin in the absence of CFTRinh172. Results CFTRinh172 significantly reduced forskolin-induced MCC but did not significantly decrease MCC-stimulated by 0.3 mM carbachol. The MCC increase to carbachol + forskolin given after either agonist alone was not inhibited by prior CFTRinh172 treatment. Conclusion The present study demonstrated an important contribution of CFTR activity to forskolin-stimulated increases in MCC. In the ex vivo ferret trachea, ∼66% inhibition was produced with CFTRinh172 pretreatment. Cholinergically stimulated MCC was reduced only ∼18% by CFTRinh172 (n.s.). MCC was synergistically increased by forskolin + carbachol. Supported by CFF and CFFT.
- Published
- 2015
32. Proteomic Analysis of Pure Human Airway Gland Mucus Reveals a Large Component of Protective Proteins
- Author
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Nam Soo Joo, Hyung Ju Cho, Idil A. Evans, Il Ho Park, Jeffrey J. Wine, and John F. Engelhardt
- Subjects
Proteomics ,Submucosal glands ,Multidisciplinary ,Innate immune system ,Mucociliary clearance ,lcsh:R ,Respiratory System ,lcsh:Medicine ,Respiratory Mucosa ,respiratory system ,Biology ,Mucus ,Cell biology ,Mucociliary Clearance ,Immunology ,Humans ,lcsh:Q ,Secretion ,Prohibitin ,lcsh:Science ,Gelsolin ,Research Article - Abstract
Airway submucosal glands contribute to innate immunity and protect the lungs by secreting mucus, which is required for mucociliary clearance and which also contains antimicrobial, anti-inflammatory, anti-proteolytic and anti-oxidant proteins. We stimulated glands in tracheal trimmings from three lung donors and collected droplets of uncontaminated mucus as they formed at the gland orifices under an oil layer. We analyzed the mucus using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Analysis identified 5486 peptides and 441 proteins from across the 3 samples (269-319 proteins per subject). We focused on 269 proteins common to at least 2 0f 3 subjects, of which 102 (38%) had protective or innate immunity functions. While many of these have long been known to play such roles, for many others their cellular protective functions have only recently been appreciated in addition to their well-studied biologic functions (e.g. annexins, apolipoproteins, gelsolin, hemoglobin, histones, keratins, and lumican). A minority of the identified proteins are known to be secreted via conventional exocytosis, suggesting that glandular secretion occurs via multiple mechanisms. Two of the observed protective proteins, major vault protein and prohibitin, have not been observed in fluid from human epithelial cultures or in fluid from nasal or bronchoalveolar lavage. Further proteomic analysis of pure gland mucus may help clarify how healthy airways maintain a sterile environment.
- Published
- 2015
33. Hyposecretion, not hyperabsorption, is the basic defect of cystic fibrosis airway glands
- Author
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Nam Soo Joo, Robert C. Robbins, Toshiya Irokawa, and Jeffrey J. Wine
- Subjects
Epithelial sodium channel ,medicine.medical_specialty ,Exocrine gland ,Time Factors ,Cystic Fibrosis ,Swine ,Cystic Fibrosis Transmembrane Conductance Regulator ,Respiratory Mucosa ,Biology ,Biochemistry ,Polymerase Chain Reaction ,Amiloride ,chemistry.chemical_compound ,Exocrine Glands ,stomatognathic system ,Chlorides ,Internal medicine ,Benzamil ,medicine ,Cyclic AMP ,Animals ,Humans ,Secretion ,Tissue Distribution ,Molecular Biology ,Lung ,DNA Primers ,Submucosal glands ,Ion Transport ,Colforsin ,Cell Biology ,respiratory system ,Mucus ,Cystic fibrosis transmembrane conductance regulator ,Trachea ,medicine.anatomical_structure ,Endocrinology ,chemistry ,biology.protein ,medicine.drug - Abstract
Human airways and glands express the anion channel cystic fibrosis transmembrane conductance regulator, CFTR, and the epithelial Na(+) channel, ENaC. Cystic fibrosis (CF) airway glands fail to secrete mucus in response to vasoactive intestinal peptide or forskolin; the failure was attributed to loss of CFTR-mediated anion and fluid secretion. Alternatively, CF glands might secrete acinar fluid via CFTR-independent pathways, but the exit of mucus from the glands could be blocked by hyperabsorption of fluid in the gland ducts. This could occur because CFTR loss can disinhibit ENaC, and ENaC activity can drive absorption. To test these two hypotheses, we measured single gland mucus secretion optically and applied ENaC inhibitors to determine whether they augmented secretion. Human CF glands were pretreated with benzamil and then stimulated with forskolin in the continued presence of benzamil. Benzamil did not rescue the lack of secretion to forskolin (50 glands, 6 CF subjects) nor did it increase the rate of cholinergically mediated mucus secretion from CF glands. Finally, neither benzamil nor amiloride increased forskolin-stimulated mucus secretion from porcine submucosal glands (75 glands, 7 pigs). One possible explanation for these results is that ENaC within the gland ducts was not active in our experiments. Consistent with that possibility, we discovered that human airway glands express Kunitz-type and non-Kunitz serine protease inhibitors, which might prevent proteolytic activation of ENaC. Our results suggest that CF glands do not display excessive, ENaC-mediated fluid absorption, leaving defective, anion-mediated fluid secretion as the most likely mechanism for defective mucus secretion from CF glands.
- Published
- 2006
34. Submucosal glands and airway defense
- Author
-
Nam Soo Joo and Jeffrey J. Wine
- Subjects
Pulmonary and Respiratory Medicine ,Respiratory Mucosa ,medicine.medical_specialty ,Exocrine gland ,Cystic Fibrosis ,Biology ,Cystic fibrosis ,fluids and secretions ,Exocrine Glands ,Internal medicine ,medicine ,Animals ,Humans ,Secretion ,Submucosal glands ,respiratory system ,medicine.disease ,Mucus ,Cystic fibrosis transmembrane conductance regulator ,Cell biology ,Serous fluid ,Endocrinology ,medicine.anatomical_structure ,biology.protein - Abstract
Most airway mucus is produced by submucosal glands in response to neural signals. Gland mucus traps microbes, inhibits their replication, and clears them from the airways. In cystic fibrosis mucus clearance is compromised, allowing pathogens to persist in static mucus. These trigger an influx of inflammatory cells, but optimal effectiveness of inflammation, and especially its resolution, also requires effective mucus clearance. Our objective is to understand the basis for defective mucus clearance in cystic fibrosis. We discovered that in subjects with cystic fibrosis, submucosal gland secretion in response to agents that elevate intracellular cyclic AMP level is completely lost and mucus stimulated by elevating intracellular Ca2+ level is thicker. We hypothesize that loss of functional cystic fibrosis transmembrane conductance regulator from gland serous cells renders them unable to secrete anions and fluid in response to any stimulus, resulting in thickened gland mucus that can be tethered to the gland ducts. In primary ciliary dyskinesias, mucus is normal, but the dysfunctional cilia lining the gland ducts may also lead to inadequate clearance of mucus from glands. Thus, understanding of lung pathology in each disease may require that an improved understanding of gland structure and function be added to our rapidly growing understanding of surface epithelia.
- Published
- 2005
35. Acid and base secretion in the Calu-3 model of human serous cells
- Author
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Mauri E. Krouse, Jeffrey J. Wine, Nam Soo Joo, Jason F. Talbott, and Martin M. Lee
- Subjects
Pulmonary and Respiratory Medicine ,medicine.medical_specialty ,Potassium Channels ,Physiology ,Respiratory Mucosa ,Biology ,Cystic fibrosis ,Models, Biological ,Cell Line ,Chlorides ,Physiology (medical) ,Internal medicine ,medicine ,Humans ,Secretion ,Submucosal glands ,Lung ,Colforsin ,Cell Biology ,Hydrogen-Ion Concentration ,medicine.disease ,Molecular biology ,Mucus ,Acetazolamide ,Serous fluid ,Bicarbonates ,medicine.anatomical_structure ,Endocrinology ,Cell culture ,Thapsigargin ,Airway - Abstract
Submucosal glands are the primary source of airway mucus, a critical component of lung innate defenses. Airway glands are defective in cystic fibrosis (CF), showing a complete absence of secretion to vasoactive intestinal peptide or forskolin, which increase intracellular cAMP concentration. This defect is attributed to gland serous cells, which express the cystic fibrosis transmembrane conductance regulator. Calu-3 cells, which mimic many features of serous cells, secrete Cl− and HCO3−, with HCO3− secretion predominating for forskolin stimulation and Cl− secretion predominating for stimuli that open basolateral K+ channels to hyperpolarize the cells. We used pH stat and ion substitution experiments to clarify the mechanisms and consequences of these two modes of secretion. We confirm that Calu-3 cells secrete primarily HCO3− in response to forskolin. Unexpectedly, HCO3− secretion continued in response to K+ channel openers, with Cl− secretion being added to it. Secretion of HCO3− from hyperpolarized cells occurs via the conversion of CO2 to HCO3− and is reduced by ∼50% with acetazolamide. A gap between the base equivalent current and short-circuit current was observed in all experiments and was traced to secretion of H+ via a ouabain-sensitive, K+-dependent process (possibly H+-K+-ATPase), which partially neutralized the secreted HCO3−. The conjoint secretion of HCO3− and H+ may help explain the puzzling finding that mucus secreted from normal and CF glands has the same acidic pH as does mucus from glands stimulated with forskolin or ACh. It may also help explain how human airway glands produce mucus that is hypotonic.
- Published
- 2004
36. A 'virtual gland' method for quantifying epithelial fluid secretion
- Author
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Nam Soo Joo, Toshiya Irokawa, Mauri E. Krouse, Jeffrey J. Wine, and Jin V. Wu
- Subjects
Pulmonary and Respiratory Medicine ,Respiratory Mucosa ,medicine.medical_specialty ,Thapsigargin ,Physiology ,Biology ,Models, Biological ,Cell Line ,chemistry.chemical_compound ,User-Computer Interface ,Physiology (medical) ,Internal medicine ,medicine ,Humans ,Secretion ,Transepithelial potential difference ,Serous cell ,Forskolin ,Colforsin ,Cell Biology ,Mucus ,Cell biology ,Kinetics ,Endocrinology ,chemistry ,Cell culture - Abstract
We developed a new apparatus, the virtual gland (VG), for measuring the rate of fluid secretion ( Jv), its composition, and the transepithelial potential (TEP) in cultured epithelial cells under open circuit. The VG creates a 10-μl chamber above the apical surface of epithelial cells on a Costar filter with a small hole leading to an oil-filled reservoir. After the chamber is primed with a fluid of choice, secreted fluid is forced through the hole into the oil, where it forms a bubble that is monitored optically to determine Jv and collected for analysis. Calu-3 cells were mounted in the VG with a basolateral bath consisting of Krebs-Ringer bicarbonate buffer at 37°C. Basal Jv was 2.7 ± 0.1 μl·cm−2·h−1 ( n = 42), and TEP was −9.2 ± 0.6 mV ( n = 33); both measures were reduced to zero by ouabain ( n = 6). Jv and TEP were stimulated 64 and 59%, respectively, by 5 μM forskolin ( n = 10), 173 and 101% by 1 mM 1-ethyl-2-benzimidazolinone ( n = 5), 213 and 122% by 333 nM thapsigargin ( n = 5), and 520 and 240% by forskolin + thapsigargin ( n = 6). Basal Jv and TEP were inhibited to 82 and 63%, respectively, with 10 μM bumetanide ( n = 5), 71 and 82% with 100 μM acetazolamide ( n = 5), and 47 and 56% with 600 μM glibenclamide ( n = 4). Basal Jv and TEP were 52 and 89% of control values, respectively, after HCO3− replacement with HEPES ( n = 16). The net HCO3− concentration of the secreted fluid was close to that of the bath (25 mM), except when stimulated with forskolin or VIP, when it increased (∼80 mM). These results validate the use of the VG apparatus and provide the first direct measures of Jv in Calu-3 cells.
- Published
- 2004
37. Mucus secretion from single submucosal glands of pig. Stimulation by carbachol and vasoactive intestinal peptide
- Author
-
Nam Soo, Joo, Yamil, Saenz, Mauri E, Krouse, and Jeffrey J, Wine
- Subjects
Swine ,Cell Membrane ,Colforsin ,Muscle, Smooth ,Hydrogen-Ion Concentration ,Trachea ,Kinetics ,Mucus ,Exocrine Glands ,Animals ,Thapsigargin ,Carbachol ,Female ,Bumetanide ,Vasoactive Intestinal Peptide - Abstract
Secretion rates of700 individual glands in isolated tracheal mucosa from 56 adult pigs were monitored optically. "Basal" secretion of 0.7 +/- 0.1 nl x min(-1) gland(-1) was observed 1-9 h post-harvest but was near zero on day 2. Secretion to carbachol (10 microm) peaked at 2-3 min and then declined to a sustained phase. Peak secretion was 12.4 +/- 1.1 nl x min(-1) gland(-1); sustained secretion was approximately one-third of peak secretion. Thapsigargin (1 microm) increased secretion from 0.1 +/- 0.05 to 0.7 +/- 0.2 nl x min(-1) gland(-1); thapsigargin did not cause contraction of the trachealis muscles. Isoproterenol and phenylephrine (10 microm each) were ineffective, but vasoactive intestinal peptide (1 microm) and forskolin (10 microm) each produced sustained secretion of 1.0 +/- 0.5 and 1.7 +/- 0.2 nl x min(-1) gland(-1), respectively. The density of actively secreting glands was 1.3/mm(2). Secretion to either carbachol or forskolin was inhibited (approximately 50%) by either bumetanide or HCO(3)(-) removal and inhibited approximately 90% by the combined treatments. Mucus secreted in response to carbachol or forskolin was acidic by approximately 0.2 pH units relative to the bath and remained acidic by approximately 0.1 pH units after bumetanide. The strong secretory response to vasoactive intestinal peptide, the acidity of [cAMP](i)-stimulated mucus, and its inhibition by bumetanide were unexpected.
- Published
- 2002
38. Submucosal gland secretions in airways from cystic fibrosis patients have normal [Na+] and pH but elevated viscosity
- Author
-
Jeffrey J. Wine, Sujatha Jayaraman, Bruce A. Reitz, Nam Soo Joo, and Alan S. Verkman
- Subjects
Exocrine gland ,medicine.medical_specialty ,Cystic Fibrosis ,medicine.medical_treatment ,Sodium ,Respiratory System ,chemistry.chemical_element ,Cystic fibrosis ,Viscosity ,Exocrine Glands ,Internal medicine ,medicine ,Humans ,Respiratory system ,Saline ,Submucosal glands ,Multidisciplinary ,respiratory system ,Biological Sciences ,Hydrogen-Ion Concentration ,medicine.disease ,Body Fluids ,medicine.anatomical_structure ,Endocrinology ,chemistry ,Biophysics ,Perfusion - Abstract
Fluid and macromolecule secretion by submucosal glands in mammalian airways is believed to be important in normal airway physiology and in the pathophysiology of cystic fibrosis (CF). Anin situfluorescence method was applied to measure the ionic composition and viscosity of freshly secreted fluid from airway glands. Fragments of human large airways obtained at the time of lung transplantation were mounted in a humidified perfusion chamber and the mucosal surface was covered by a thin layer of oil. Individual droplets of secreted fluid were microinjected with fluorescent indicators for measurement of [Na+], [Cl−], and pH by ratio imaging fluorescence microscopy and viscosity by fluorescence recovery after photobleaching. After carbachol stimulation, 0.1–0.5 μl of fluid accumulated in spherical droplets at gland orifices in ≈3–5 min. In gland fluid from normal human airways, [Na+] was 94 ± 8 mM, [Cl−] was 92 ± 12 mM, and pH was 6.97 ± 0.06 (SE,n= 7 humans, more than five glands studied per sample). Apparent fluid viscosity was 2.7 ± 0.3-fold greater than that of saline. Neither [Na+] nor pH differed in gland fluid from CF airways, but viscosity was significantly elevated by ≈2-fold compared to normal airways. These results represent the first direct measurements of ionic composition and viscosity in uncontaminated human gland secretions and indicate similar [Na+], [Cl−], and pH to that in the airway surface liquid. The elevated gland fluid viscosity in CF may be an important factor promoting bacterial colonization and airway disease.
- Published
- 2001
39. QUEST FOR CYSTIC FIBROSIS PHARMACOTHERAPY
- Author
-
Nam Soo Joo
- Subjects
Pharmacology ,Pediatrics ,medicine.medical_specialty ,Pharmacotherapy ,Physiology ,business.industry ,Physiology (medical) ,Medicine ,business ,medicine.disease ,Cystic fibrosis - Published
- 2008
40. Regulation of intestinal Cl- and HCO3-secretion by uroguanylin
- Author
-
Leonard R. Forte, Lane L. Clarke, Roslyn M. London, Hyun Dju Kim, and Nam Soo Joo
- Subjects
Anions ,medicine.medical_specialty ,Physiology ,Duodenum ,Guanylin ,Peptide hormone ,chemistry.chemical_compound ,Mice ,Chlorides ,Physiology (medical) ,Internal medicine ,medicine ,Animals ,Secretion ,Intestinal Mucosa ,Receptor ,Natriuretic Peptides ,Cecum ,Ion transporter ,Hepatology ,Gastroenterology ,Electric Conductivity ,Bicarbonate transport ,Hydrogen-Ion Concentration ,Epithelium ,Intestines ,Mice, Inbred C57BL ,Bicarbonates ,Endocrinology ,medicine.anatomical_structure ,chemistry ,Female ,Peptides ,Uroguanylin - Abstract
Uroguanylin is an intestinal peptide hormone that may regulate epithelial ion transport by activating a receptor guanylyl cyclase on the luminal surface of the intestine. In this study, we examined the action of uroguanylin on anion transport in different segments of freshly excised mouse intestine, using voltage-clamped Ussing chambers. Uroguanylin induced larger increases in short-circuit current (Isc) in proximal duodenum and cecum compared with jejunum, ileum, and distal colon. The acidification of the lumen of the proximal duodenum (pH 5.0-5.5) enhanced the stimulatory action of uroguanylin. In physiological Ringer solution, a significant fraction of the Isc stimulated by uroguanylin was insensitive to bumetanide and dependent on HCO3- in the bathing medium. Experiments using pH-stat titration revealed that uroguanylin stimulates serosal-to-luminal HCO3- secretion (Js--lHCO3-) together with a larger increase in Isc. Both Js--lHCO3- and Isc were significantly augmented when luminal pH was reduced to pH 5.15. Uroguanylin also stimulated the Js--lHCO3- and Isc across the cecum, but luminal acidity caused a generalized decrease in the bioelectric responsiveness to agonist stimulation. In cystic fibrosis transmembrane conductance regulator (CFTR) knockout mice, the duodenal Isc response to uroguanylin was markedly reduced, but not eliminated, despite having a similar density of functional receptors. It was concluded that uroguanylin is most effective in acidic regions of the small intestine, where it stimulates both HCO3- and Cl-secretion primarily via a CFTR-dependent mechanisms.
- Published
- 1998
41. In Vivo Readout of CFTR Function: Ratiometric Measurement of CFTR-Dependent Secretion by Individual, Identifiable Human Sweat Glands
- Author
-
Sara E. Modlin, Jonathan H. Chen, Hyung Ju Cho, Jessica E. Char, Carlos Milla, Nam Soo Joo, Kim V. Tran, Il Ho Park, Jeffrey J. Wine, Rohan Verma, Marlene H. Wolfe, Ewart A. C. Thomas, Eric Frisbee, and Colleen Dunn
- Subjects
Adult ,Atropine ,Male ,Heterozygote ,medicine.medical_specialty ,Time Factors ,Cystic Fibrosis ,Injections, Intradermal ,Cystic Fibrosis Transmembrane Conductance Regulator ,lcsh:Medicine ,Muscarinic Antagonists ,Muscarinic Agonists ,Cystic fibrosis ,In vivo ,Internal medicine ,Muscarinic acetylcholine receptor ,medicine ,Humans ,Bioassay ,Secretion ,Sweat ,lcsh:Science ,Methacholine Chloride ,Multidisciplinary ,Dose-Response Relationship, Drug ,integumentary system ,biology ,lcsh:R ,Isoproterenol ,Heterozygote advantage ,Adrenergic beta-Agonists ,medicine.disease ,Aminophylline ,Cystic fibrosis transmembrane conductance regulator ,Sweat Glands ,Endocrinology ,Purinergic P1 Receptor Antagonists ,Mutation ,biology.protein ,Female ,lcsh:Q ,Methacholine ,Research Article ,medicine.drug - Abstract
To assess CFTR function in vivo, we developed a bioassay that monitors and compares CFTR-dependent and CFTR-independent sweat secretion in parallel for multiple (~50) individual, identified glands in each subject. Sweating was stimulated by intradermally injected agonists and quantified by optically measuring spherical sweat bubbles in an oil-layer that contained dispersed, water soluble dye particles that partitioned into the sweat bubbles, making them highly visible. CFTR-independent secretion (M-sweat) was stimulated with methacholine, which binds to muscarinic receptors and elevates cytosolic calcium. CFTR-dependent secretion (C-sweat) was stimulated with a β-adrenergic cocktail that elevates cytosolic cAMP while blocking muscarinic receptors. A C-sweat/M-sweat ratio was determined on a gland-by-gland basis to compensate for differences unrelated to CFTR function, such as gland size. The average ratio provides an approximately linear readout of CFTR function: the heterozygote ratio is ~0.5 the control ratio and for CF subjects the ratio is zero. During assay development, we measured C/M ratios in 6 healthy controls, 4 CF heterozygotes, 18 CF subjects and 4 subjects with 'CFTR-related' conditions. The assay discriminated all groups clearly. It also revealed consistent differences in the C/M ratio among subjects within groups. We hypothesize that these differences reflect, at least in part, levels of CFTR expression, which are known to vary widely. When C-sweat rates become very low the C/M ratio also tended to decrease; we hypothesize that this nonlinearity reflects ductal fluid absorption. We also discovered that M-sweating potentiates the subsequent C-sweat response. We then used potentiation as a surrogate for drugs that can increase CFTR-dependent secretion. This bioassay provides an additional method for assessing CFTR function in vivo, and is well suited for within-subject tests of systemic, CFTR-directed therapeutics.
- Published
- 2013
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