Purification and biochemical characterization of pulmonary surfactant protein A of horses.
Abstract: To characterize surfactant protein isolated from bronchoalveolar lavage fluids of healthy horses. Methods: 10 Thoroughbreds (5 males, 5 females; 26 to 40 months old) that did not have a history or clinical signs of respiratory tract disease. Methods: Bronchoalveolar lavage fluid (BALF) was obtained and centrifuged at 33,000 X g. Lipid was removed from precipitated fractions by means of extraction with 1-butanol, and organic solvent-insoluble protein precipitates were dialyzed against Tris buffer. The suspension was centrifuged, and supernatant was placed in a mannose-Sepharose affinity column, with calcium. The bound protein fraction was analyzed by means of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, western immunoblot analysis, and amino acid sequencing. A liposome-aggregation assay was also performed, using purified proteins. Results: Protein isolated by use of mannose-affinity matrices was identified as surfactant protein A (SP-A). It had carbohydrate-binding and phospholipid-aggregation properties characteristic of SP-A isolated from other animal species. The partial primary sequence of the isolated protein had high homology with rat and human SP-A. Furthermore, the equine SP-A reacted with anti-human and anti-rat SP-A specific antibodies. Conclusions: Analysis of these findings indicated the existence of SP-A in pulmonary tissues of horses. Conclusions: Measurement of SP-A concentrations may be useful for clinicians evaluating pulmonary disease of horses.
Publication Date: 1999-02-27 PubMed ID: 10048546
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- Journal Article
- Amino Acid Sequence
- Animal Species
- Biochemistry
- Bronchoalveolar Lavage
- Clinical Pathology
- Comparative Study
- Diagnosis
- Diagnostic Technique
- Disease Diagnosis
- Equine Diseases
- Equine Health
- Horses
- Immunology
- Laboratory Methods
- Protein
- Pulmonary Health
- Respiratory Disease
- Thoroughbreds
- Veterinary Medicine
- Veterinary Research
- Western Blot
Summary
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This study focuses on the isolation and characterization of surfactant protein A (SP-A) from the lung fluids of healthy horses. This protein was found to have characteristics similar to SP-A from other species and its measurement could be beneficial for veterinary clinicians assessing horses’ lung diseases.
Background
The article seeks to isolate and examine the characteristics of surfactant protein A (SP-A) found in bronchoalveolar lavage fluid (BALF), a substance collected from the lungs of healthy horses. SurfActant proteins are important in lung function because they reduce surface tension and prevent the lungs from collapsing. Studying their properties could lead to better understanding and treatment of pulmonary diseases in horses.
Methods
- The research subjects include 10 healthy thoroughbred horses, without any traces or symptoms of respiratory diseases.
- BALF was extracted from these horses and centrifuged at high speed. The resulting precipitate was cleaned of lipids through 1-butanol extraction.
- The resulting insoluble protein was then dialyzed and centrifuged. The supernatant was processed using a mannose-Sepharose affinity column to isolate certain proteins, with calcium present.
- The end product was scrutinized using sodium dodecyl sulfate-polyacrylamide gel electrophoresis, Western immunoblot analysis, and amino acid sequencing. Additionally, a liposome-aggregation assay was performed using the purified proteins.
Results
- The protein isolated via the mannose-affinity matrices was identified as surfactant protein A (SP-A), displaying carbohydrate-binding and phospholipid-aggregation properties, which are shared by SP-As of other species.
- The partial primary sequence of the equine SP-A showed a high degree of similarity with rat and human SP-As. Moreover, the horse-derived SP-A reacted with anti-human and anti-rat SP-A specific antibodies.
Conclusions
- The study confirmed the presence of SP-A in the horse’s lung tissue, broadening the understanding of equine pulmonary health.
- The ability to measure SP-A concentrations could provide veterinary practitioners with a valuable tool for examining and diagnosing pulmonary diseases in horses.
Cite This Article
APA
Hobo S, Ogasawara Y, Kuroki Y, Akino T, Yoshihara T.
(1999).
Purification and biochemical characterization of pulmonary surfactant protein A of horses.
Am J Vet Res, 60(2), 169-173.
Publication
Researcher Affiliations
- Clinical Science and Pathobiology Division, Equine Research Institute, Japan Racing Association, Tochigi.
MeSH Terms
- Amidohydrolases / chemistry
- Amino Acid Sequence
- Amino Acids / analysis
- Animals
- Antibodies, Monoclonal
- Blotting, Western / veterinary
- Bronchoalveolar Lavage / veterinary
- Bronchoalveolar Lavage Fluid / chemistry
- Chromatography, Agarose / veterinary
- Collagenases / chemistry
- Electrophoresis, Polyacrylamide Gel / veterinary
- Female
- Horses / physiology
- Liposomes / physiology
- Lung / chemistry
- Lung / physiology
- Male
- Molecular Sequence Data
- Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
- Phospholipids / chemistry
- Proteolipids / chemistry
- Proteolipids / isolation & purification
- Proteolipids / physiology
- Pulmonary Surfactant-Associated Protein A
- Pulmonary Surfactant-Associated Proteins
- Pulmonary Surfactants / chemistry
- Pulmonary Surfactants / isolation & purification
- Pulmonary Surfactants / physiology
- Sequence Analysis
- Sequence Homology, Amino Acid
Citations
This article has been cited 3 times.- Bocking T, Singh B. Light and electron-microscopic localization of CD9 and surfactant protein A and D in normal lungs of the horse. Can J Vet Res 2021 Jul;85(3):170-176.
- Yamaya Y, Suzuki K, Watari T, Asano R. Bronchoalveolar lavage fluid and serum canine surfactant protein A concentrations in dogs with chronic cough by bronchial and interstitial lung diseases. J Vet Med Sci 2014 Apr;76(4):593-6.
- Kankavi O, Roberts MS. Detection of surfactant protein A (SP-A) and surfactant protein D (SP-D) in equine synovial fluid with immunoblotting. Can J Vet Res 2004 Apr;68(2):146-9.
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