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Quantification of mucin gene expression in tracheobronchial epithelium of healthy dogs and dogs with chronic bronchitis

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  • 1 Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27606.
  • | 2 Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27606.
  • | 3 Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27606.
  • | 4 Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27606.
  • | 5 Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27606.
  • | 6 Department of Molecular Biomedical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27606.

Abstract

Objective—To develop a real-time PCR assay for the quantification of mucin gene expression in tracheobronchial brushing specimens from dogs and compare mucin gene expression in specimens from dogs with naturally occurring chronic bronchitis with that in specimens from healthy dogs.

Animals—7 healthy dogs and 5 dogs with chronic bronchitis.

Procedures—Primers that were designed to span the predicted intron-exon boundaries of a canine MUC5AC-like gene were used to develop a real-time PCR assay for quantification of expression of that gene. Total mRNA was isolated from tracheobronchial brushing specimens obtained from dogs with and without bronchitis during anesthesia; MUC5AC-like gene expression in those samples was quantified by use of the real-time PCR assay.

Results—The PCR assay was sensitive and specific for the target sequence, the predicted amino acid sequence of which had greatest homology with human, porcine, and rat MUC5AC. The assay was able to quantify the target over a wide dynamic range. Dogs with chronic bronchitis had a 3.0-fold increase in the quantity of MUC5AC-like mRNA, compared with healthy dogs.

Conclusions and Clinical Relevance—The ability to measure mucin gene expression from tracheobronchial brushing specimens collected from client-owned dogs during routine bronchoscopy should prove to be a useful tool for the study of bronchitis in dogs and expand the usefulness of airway inflammation in dogs as a model for bronchitis in humans.

Abstract

Objective—To develop a real-time PCR assay for the quantification of mucin gene expression in tracheobronchial brushing specimens from dogs and compare mucin gene expression in specimens from dogs with naturally occurring chronic bronchitis with that in specimens from healthy dogs.

Animals—7 healthy dogs and 5 dogs with chronic bronchitis.

Procedures—Primers that were designed to span the predicted intron-exon boundaries of a canine MUC5AC-like gene were used to develop a real-time PCR assay for quantification of expression of that gene. Total mRNA was isolated from tracheobronchial brushing specimens obtained from dogs with and without bronchitis during anesthesia; MUC5AC-like gene expression in those samples was quantified by use of the real-time PCR assay.

Results—The PCR assay was sensitive and specific for the target sequence, the predicted amino acid sequence of which had greatest homology with human, porcine, and rat MUC5AC. The assay was able to quantify the target over a wide dynamic range. Dogs with chronic bronchitis had a 3.0-fold increase in the quantity of MUC5AC-like mRNA, compared with healthy dogs.

Conclusions and Clinical Relevance—The ability to measure mucin gene expression from tracheobronchial brushing specimens collected from client-owned dogs during routine bronchoscopy should prove to be a useful tool for the study of bronchitis in dogs and expand the usefulness of airway inflammation in dogs as a model for bronchitis in humans.

Contributor Notes

Mr. Large's present address is Department of Genetics, College of Agricultural and Life Sciences, North Carolina State University, Raleigh, NC 27606.

Supported by grants from the Morris Animal Foundation through sponsorship by Fort Worth Tarrant County Animal Foundation; the North Carolina State University Competitive grants program; the Department of Clinical Sciences, North Carolina State University; and grant R37HL36982 from the National Institutes of Health.

Address correspondence to Dr. Hawkins.