Identification of cystatin C, a cysteine proteinase inhibitor, as a major secretory product of human alveolar macrophages in vitro
Chapman Jr., Harold A. ; Reilly Jr., John J. ; Yee, Robert and Grubb, Anders LU
(1990)
In The American Review of Respiratory Disease
141(3).
p.698-705
- Abstract
The major inhibitor of the cysteine class of proteinases found in human body fluids, such as spinal fluid, milk, and seminal plasma, is cystatin C. In this study we show that human bronchoalveolar fluid also contains cystatin C and examine cystatin C expression by alveolar macrophages in vitro. Immunoprecipitation of extracts of metabolically labeled cells and immunoblotting of cellular extracts and culture media show that cystatin C is synthesized as a 14 (+/- 0.5) kilodalton (kD) protein and that greater than 90% of the protein is released as the 14 kD product into the culture supernatant (26.5 +/- 6.8 ng per 10(6) cells per 24 h). Cystatin C is not one of the most abundant proteins secreted during the first 24 h in vitro,... (More)
The major inhibitor of the cysteine class of proteinases found in human body fluids, such as spinal fluid, milk, and seminal plasma, is cystatin C. In this study we show that human bronchoalveolar fluid also contains cystatin C and examine cystatin C expression by alveolar macrophages in vitro. Immunoprecipitation of extracts of metabolically labeled cells and immunoblotting of cellular extracts and culture media show that cystatin C is synthesized as a 14 (+/- 0.5) kilodalton (kD) protein and that greater than 90% of the protein is released as the 14 kD product into the culture supernatant (26.5 +/- 6.8 ng per 10(6) cells per 24 h). Cystatin C is not one of the most abundant proteins secreted during the first 24 h in vitro, representing approximately 10 to 12% of the total protein released by normal nonsmoker macrophages. Alveolar macrophages obtained from cigarette smokers or nonsmoker macrophages exposed to zymosan in vitro released 10 to 55% less cystatin C than nonsmoker macrophages. We also assayed culture supernatants from macrophages of smokers and nonsmokers for functional cystatin C. Supernatants of nonsmoker macrophages inhibited cathepsin B-like amidolytic activity in a fluorometric assay at pH 5.5. The inhibition was blocked by adsorption with Sepharose-coupled cystatin C antibodies and the inhibitor subsequently recovered from the Sepharose beads. In contrast, supernatants from smoker macrophages had obvious cathepsin B-like activity.(ABSTRACT TRUNCATED AT 250 WORDS)
(Less)
- author
- Chapman Jr., Harold A.
; Reilly Jr., John J.
; Yee, Robert
and Grubb, Anders
LU
- organization
- publishing date
- 1990
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- Bronchoalveolar Lavage Fluid/metabolism, Cathepsins/antagonists & inhibitors, Cells, Cultured, Cystatin C, Cystatins/biosynthesis, Cysteine Proteinase Inhibitors/metabolism, Humans, Inflammation, Macrophages/metabolism, Proteins/metabolism, Pulmonary Alveoli/cytology, Smoking/metabolism, Zymosan/pharmacology
- in
- The American Review of Respiratory Disease
- volume
- 141
- issue
- 3
- pages
- 698 - 705
- publisher
- American Thoracic Society
- external identifiers
-
- scopus:0025320213
- pmid:2310099
- ISSN
- 0003-0805
- DOI
- 10.1164/ajrccm/141.3.698
- language
- English
- LU publication?
- yes
- id
- 2006544a-aabe-4f49-b169-320741156540
- date added to LUP
- 2021-10-27 13:15:45
- date last changed
- 2024-01-12 02:54:30
@article{2006544a-aabe-4f49-b169-320741156540,
abstract = {{<p>The major inhibitor of the cysteine class of proteinases found in human body fluids, such as spinal fluid, milk, and seminal plasma, is cystatin C. In this study we show that human bronchoalveolar fluid also contains cystatin C and examine cystatin C expression by alveolar macrophages in vitro. Immunoprecipitation of extracts of metabolically labeled cells and immunoblotting of cellular extracts and culture media show that cystatin C is synthesized as a 14 (+/- 0.5) kilodalton (kD) protein and that greater than 90% of the protein is released as the 14 kD product into the culture supernatant (26.5 +/- 6.8 ng per 10(6) cells per 24 h). Cystatin C is not one of the most abundant proteins secreted during the first 24 h in vitro, representing approximately 10 to 12% of the total protein released by normal nonsmoker macrophages. Alveolar macrophages obtained from cigarette smokers or nonsmoker macrophages exposed to zymosan in vitro released 10 to 55% less cystatin C than nonsmoker macrophages. We also assayed culture supernatants from macrophages of smokers and nonsmokers for functional cystatin C. Supernatants of nonsmoker macrophages inhibited cathepsin B-like amidolytic activity in a fluorometric assay at pH 5.5. The inhibition was blocked by adsorption with Sepharose-coupled cystatin C antibodies and the inhibitor subsequently recovered from the Sepharose beads. In contrast, supernatants from smoker macrophages had obvious cathepsin B-like activity.(ABSTRACT TRUNCATED AT 250 WORDS)</p>}},
author = {{Chapman Jr., Harold A. and Reilly Jr., John J. and Yee, Robert and Grubb, Anders}},
issn = {{0003-0805}},
keywords = {{Bronchoalveolar Lavage Fluid/metabolism; Cathepsins/antagonists & inhibitors; Cells, Cultured; Cystatin C; Cystatins/biosynthesis; Cysteine Proteinase Inhibitors/metabolism; Humans; Inflammation; Macrophages/metabolism; Proteins/metabolism; Pulmonary Alveoli/cytology; Smoking/metabolism; Zymosan/pharmacology}},
language = {{eng}},
number = {{3}},
pages = {{698--705}},
publisher = {{American Thoracic Society}},
series = {{The American Review of Respiratory Disease}},
title = {{Identification of cystatin C, a cysteine proteinase inhibitor, as a major secretory product of human alveolar macrophages in vitro}},
url = {{http://dx.doi.org/10.1164/ajrccm/141.3.698}},
doi = {{10.1164/ajrccm/141.3.698}},
volume = {{141}},
year = {{1990}},
}