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Canadian journal of veterinary research = Revue canadienne de recherche veterinaire2008; 72(4); 340-349;

Characterization of endothelin receptors in the peripheral lung tissues of horses unaffected and affected with recurrent airway obstruction.

Abstract: The purpose of the study was to determine and compare the expression of endothelin (ET) receptors in the peripheral lungs of healthy horses and those affected with recurrent airway obstruction (RAO) using reverse transcriptase polymerase chain reaction (RT-PCR), real-time PCR, Western blot analysis, and immunohistochemical techniques. Two groups of horses (7 healthy and 7 RAO-affected) were selected from a pool of horses destined for euthanasia. The grouping of horses was based on the history, clinical scoring, and pulmonary function testing. After euthanasia, gross postmortem evaluation of the lungs was conducted, and lung samples were collected and either stored at -80 degrees C or fixed in zinc-formalin for 12 h. The RT-PCR was performed by using specific primers for ETA and ETB receptors, and beta-actin. To determine the relative gene expression real-time PCR was performed. To detect ET receptor protein expression, Western blotting and immunohistochemical studies were performed using polyclonal antibodies against ETA and ETB receptors and beta-actin. The ET receptor expression was determined by performing either densitometric analyses or scoring of immunostaining. Statistical analyses were performed to detect differences in receptor expression within and between the 2 groups. The results indicated that ET receptor expression, particularly ETB receptors, was significantly greater in the peripheral lungs of RAO-affected horses than in those of healthy horses. Clinical trials using ET receptor antagonists, particularly ETB antagonists might help in developing a therapeutic strategy to treat this career-ending disease. L’objectif de la présente étude était de déterminer et comparer l’expression des récepteurs de l’endothéline (ET) dans les poumons périphériques de chevaux en santé et de chevaux souffrant d’obstruction récurrente des voies respiratoires (RAO) par réaction d’amplification en chaîne par la polymérase à l’aide de la transcriptase réverse (RT-PCR), par PCR en temps réel, par réaction d’immunobuvardage et par techniques immunohistochimiques. Deux groupes de chevaux (7 en santé et 7 atteints de RAO) ont été sélectionnés à partir d’un regroupement de chevaux destinés à l’euthanasie. Le groupement des chevaux s’est fait sur la base de l’anamnèse, le pointage clinique, et un test de fonction respiratoire. Après l’euthanasie, une évaluation des lésions macroscopiques des poumons a été faite, et des échantillons de poumons prélevés et soit entreposés à −80 °C ou fixé pendant 12 h dans une solution de zinc-formaline. L’épreuve de RT-PCR a été effectuée en utilisant des amorces spécifiques pour les récepteurs ETA et ETB, et β-actine. Afin de déterminer l’expression génique relative une épreuve de PCR en temps réel a été effectuée. Afin de déterminer l’expression des récepteurs ET, les épreuves d’immunobuvardage et immunohistochimiques ont été effectuées en utilisant des anticorps polyclonaux contre les récepteurs d’ETA et d’ETB ainsi que ceux pour β-actine. L’expression de récepteur ET a été déterminée en réalisant des analyses densitométriques ou en attribuant un pointage à l’immunocoloration. Les analyses statistiques ont été faites afin de détecter des différences dans l’expression des récepteurs à l’intérieur et entre les 2 groupes. Les résultats indiquent que l’expression des récepteurs ET, en particulier les récepteurs ETB, était significativement plus grande dans les poumons périphériques des chevaux atteints de RAO que ceux des chevaux en santé. Des essais cliniques utilisant des antagonistes des récepteurs ET, plus particulièrement les antagonistes ETB, pourraient aider au développement d’une stratégie pour traiter cette maladie qui met fin à une carrière. (Traduit par Docteur Serge Messier)
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Publication Date: 2008-09-12 PubMed ID: 18783023PubMed Central: PMC2442677
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  • Journal Article
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  • Non-U.S. Gov't
  • Research Support
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  • Non-P.H.S.

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This study aimed to evaluate and contrast the presence of endothelin receptors in the lungs of both healthy horses and horses suffering from recurrent airway obstruction (RAO). The test results indicated that horses with RAO have a significantly higher expression of these receptors, particularly ETB receptors, compared to healthy horses, suggesting ET receptor antagonists could be beneficial in treating this ailment.

Objective and Methodology

  • The study was designed to understand the expression of Endothelin (ET) receptors in the peripheral lungs of healthy horses and those with recurrent airway obstruction (RAO).
  • Fourteen horses were selected for the study, seven of which were healthy and seven of which were suffering from RAO. Selection was based on their medical history, clinical scoring, and pulmonary function.
  • The horses were eventually euthanized, and postmortem examination of the lungs was conducted to collect lung samples.
  • The lung samples were stored at extreme low temperatures or fixed in zinc-formalin for 12 hours.

Experimental Procedures

  • Reverse transcriptase polymerase chain reaction (RT-PCR), real-time PCR, Western blot analysis, and immunohistochemical techniques were used in the study. These are standard methods to detect and quantify specific genes and proteins in tissue samples.
  • For RT-PCR, specific primers for ETA and ETB receptors, and beta-actin were used.
  • Real-time PCR was performed to determine the relative gene expression. Western blotting and immunohistochemical studies were performed to detect ET receptor protein expression.
  • The studies used polyclonal antibodies against ETA and ETB receptors and beta-actin.
  • The density of the receptor expression was determined either by densitometric analyses or by scoring the immunostaining.
  • Statistical analysis was used to detect and quantify differences in receptor expression both within the same group of horses and between the two groups.

Findings and Implications

  • Results showed that the expression of ET receptors, especially ETB receptors, was significantly higher in RAO-affected horses than in healthy horses.
  • This discovery implies that these receptors might play a significant role in the development or persistence of RAO in horses.
  • The findings suggest that treating RAO-affected horses with ET receptor antagonists, particularly ETB antagonists, might help in controlling this debilitating disease.

Cite This Article

APA
Polikepahad S, Haque M, Francis J, Moore RM, Venugopal CS. (2008). Characterization of endothelin receptors in the peripheral lung tissues of horses unaffected and affected with recurrent airway obstruction. Can J Vet Res, 72(4), 340-349.

Publication

Canadian journal of veterinary research = Revue canadienne de recherche veterinaire
ISSN: 0830-9000
NlmUniqueID: 8607793
Country: Canada
Language: English
Volume: 72
Issue: 4
Pages: 340-349

Researcher Affiliations

Polikepahad, Sumanth
  • Equine Health Studies Program, Department of Comparative Biomedical Sciences, Louisiana State University, Baton Rouge, Louisiana, 70803 USA.
Haque, Masudul
    Francis, Joseph
      Moore, Rustin M
        Venugopal, Changaram S

          MeSH Terms

          • Animals
          • Blotting, Western / veterinary
          • Case-Control Studies
          • Densitometry
          • Gene Expression
          • Horse Diseases / metabolism
          • Horses
          • Immunohistochemistry / veterinary
          • Lung / metabolism
          • Lung / pathology
          • Lung Diseases, Obstructive / metabolism
          • Lung Diseases, Obstructive / veterinary
          • Polymerase Chain Reaction / veterinary
          • Receptors, Endothelin / genetics
          • Receptors, Endothelin / metabolism
          • Reverse Transcriptase Polymerase Chain Reaction / veterinary

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