Effect of chondroitinase ABC on purulent sputum from cystic fibrosis and other patients.
Cystic fibrosis (CF) patients develop chronic lung infections associated with airway obstruction by viscous and insoluble mucus secretions. Although mucus glycoproteins (mucins) are thought to be responsible for mucus plugs, other glycoconjugate components of airway secretions have not been systematically evaluated. The aim of the present study was to determine whether chondroitin sulfate proteoglycans (CSPG) contribute to the insolubility of CF sputum. Sputa obtained from 18 CF patients were incubated with chondroitinase ABC (ChABC) or buffer (control) for 18 h at 37 degrees C, and after centrifugation at 12,000 g, the volume of the insoluble pellet and turbidity of the supernatant were determined as measures of solubility. ChABC caused a 70-90% reduction in supernatant turbidity and a 60-70% decrease in pellet volume of the 13 purulent CF sputa, but had much less effect on the five nonpurulent CF sputa tested. Similar results were obtained with two non-CF purulent and two non-CF, nonpurulent sputa. Gel electrophoresis, Western blot, and slot blot immunoassays with antichondroitin sulfate and antimucin antibodies revealed that purulent sputa (CF and non-CF) contained more CSPG and less mucin than nonpurulent sputa. In vitro mixing experiments showed that mucin in nonpurulent sputa was reduced upon incubation with purulent sputa, presumably because of degradation or a loss of immunoreactive mucin epitopes from leukocyte and/or bacterial enzymes present in purulent sputa. Our results suggest that CSPG contribute more significantly than mucins to the insolubility of purulent tracheobronchial secretions from CF patients. Because purulent sputa from non-CF patients showed a similar pattern, our observations with CF sputa may have wider applicability.
Khatri
Ismat A
IA
Research Institute, Department of Structural Biology and Biochemistry, The Hospital for Sick Children, Toronto, Ontario, Canada.
Bhaskar
K Ramakrishnan
KR
Lamont
J Thomas
JT
Sajjan
S Umadevi
SU
Ho
Catherine K Y
CK
Forstner
Janet
J
D000328:Adult; D000906:Antibodies / Q000494:pharmacology; D015153:Blotting, Western; D019765:Chondroitin ABC Lyase / Q000494:pharmacology*; D011508:Chondroitin Sulfate Proteoglycans / Q000302:isolation & purification / Q000378:metabolism*; D003550:Cystic Fibrosis / Q000378:metabolism*; D004588:Electrophoresis, Cellulose Acetate; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D009077:Mucins / Q000276:immunology / Q000378:metabolism; D012995:Solubility; D013183:Sputum / Q000187:drug effects* / Q000378:metabolism*
D000906:Antibodies; D011508:Chondroitin Sulfate Proteoglycans; D009077:Mucins; D019765:Chondroitin ABC Lyase