title: No evidence for altered intracellular calcium-handling in airway smooth muscle cells from human subjects with asthma.
abstract: BACKGROUND
  Asthma is characterized by airway hyper-responsiveness and variable airflow obstruction, in part as a consequence of hyper-contractile airway smooth muscle, which persists in primary cell culture. One potential mechanism for this hyper-contractility is abnormal intracellular Ca(2+) handling.
  
  
  METHODS
  We sought to compare intracellular Ca(2+) handling in airway smooth muscle cells from subjects with asthma compared to non-asthmatic controls by measuring: i) bradykinin-stimulated changes in inositol 1,4,5-trisphosphate (IP3) accumulation and intracellular Ca(2+) concentration, ii) sarco/endoplasmic reticulum Ca(2+)-ATPase (SERCA) expression, iii) mechanisms of cytoplasmic Ca(2+) clearance assessed following instantaneous flash photolytic release of Ca(2+) into the cytoplasm.
  
  
  RESULTS
  We found no differences in airway smooth muscle cell basal intracellular Ca(2+) concentrations, bradykinin-stimulated IP3 accumulation or intracellular Ca(2+) responses. Quantification of SERCA2 mRNA or protein expression levels revealed no differences in ASM cells obtained from subjects with asthma compared to non-asthmatic controls. We did not identify differences in intracellular calcium kinetics assessed by flash photolysis and calcium uncaging independent of agonist-activation with or without SERCA inhibition. However, we did observe some correlations in subjects with asthma between lung function and the different cellular measurements of intracellular Ca(2+) handling, with poorer lung function related to increased rate of recovery following flash photolytic elevation of cytoplasmic Ca(2+) concentration.
  
  
  CONCLUSIONS
  Taken together, the experimental results reported in this study do not demonstrate major fundamental differences in Ca(2+) handling between airway smooth muscle cells from non-asthmatic and asthmatic subjects. Therefore, increased contraction of airway smooth muscle cells derived from asthmatic subjects cannot be fully explained by altered Ca(2+) homeostasis.
authors.affiliation: Department of Infection, Immunity & Inflammation, and Institute for Lung Health, University of Leicester, Glenfield Hospital, Leicester, LE3 9QP, UK. davidsweeney111@gmail.com.|Department of Cell Physiology & Pharmacology, University of Leicester, Henry Wellcome Building, Lancaster Road, Leicester, LE1 9HN, UK. davidsweeney111@gmail.com.
authors.forename: David
authors.initials: D
authors.lastname: Sweeney
authors.affiliation: Department of Infection, Immunity & Inflammation, and Institute for Lung Health, University of Leicester, Glenfield Hospital, Leicester, LE3 9QP, UK. fmh7@le.ac.uk.
authors.forename: Fay
authors.initials: F
authors.lastname: Hollins
authors.affiliation: Department of Infection, Immunity & Inflammation, and Institute for Lung Health, University of Leicester, Glenfield Hospital, Leicester, LE3 9QP, UK. eg32@le.ac.uk.|Department of Cell Physiology & Pharmacology, University of Leicester, Henry Wellcome Building, Lancaster Road, Leicester, LE1 9HN, UK. eg32@le.ac.uk.
authors.forename: Edith
authors.initials: E
authors.lastname: Gomez
authors.affiliation: Department of Cell Physiology & Pharmacology, University of Leicester, Henry Wellcome Building, Lancaster Road, Leicester, LE1 9HN, UK. mis@le.ac.uk.
authors.forename: Rajendra
authors.initials: R
authors.lastname: Mistry
authors.affiliation: Department of Infection, Immunity & Inflammation, and Institute for Lung Health, University of Leicester, Glenfield Hospital, Leicester, LE3 9QP, UK. rms4@le.ac.uk.
authors.forename: Ruth
authors.initials: R
authors.lastname: Saunders
authors.affiliation: Department of Cell Physiology & Pharmacology, University of Leicester, Henry Wellcome Building, Lancaster Road, Leicester, LE1 9HN, UK. jc36@leicester.ac.uk.
authors.forename: Robert Alfred John
authors.initials: RA
authors.lastname: Challiss
authors.affiliation: Department of Infection, Immunity & Inflammation, and Institute for Lung Health, University of Leicester, Glenfield Hospital, Leicester, LE3 9QP, UK. ceb17@leicester.ac.uk.
authors.forename: Christopher Edward
authors.initials: CE
authors.lastname: Brightling
mesh_terms: D000328:Adult; D001249:Asthma / Q000235:genetics / Q000378:metabolism*; D001920:Bradykinin / Q000494:pharmacology; D001980:Bronchi / Q000166:cytology; D002118:Calcium / Q000378:metabolism*; D016022:Case-Control Studies; D005260:Female; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D009119:Muscle Contraction; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000378:metabolism*; D010782:Photolysis; D012333:RNA, Messenger / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000235:genetics* / Q000378:metabolism; D014665:Vasodilator Agents / Q000494:pharmacology
chemical_list: D012333:RNA, Messenger; D014665:Vasodilator Agents; D015544:Inositol 1,4,5-Trisphosphate; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; C506946:ATP2A2 protein, human; D001920:Bradykinin; D002118:Calcium