Publication: Obtención de proteínas de membrana externa de bacterias periodontopatógenas y análisis de la respuesta inmunitaria de pacientes con periodontitis
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2018
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Introducción: Las enfermedades periodontales son el resultado de la acumulación de microorganismos alrededor del diente y de la respuesta inmunitaria asociada. Las alteraciones provocadas por los mecanismos de defensa, en un intento de frenar el avance bacteriano, conducen a la pérdida de los tejidos de soporte. El cambio a una microbiota más patogénica, gram-negativa anaeróbica, dominada por microorganismos periodontopatógenos, puede marcar el inicio de la pérdida de inserción. Objetivo: El propósito de este estudio ha sido comparar la respuesta inmunitaria de pacientes con periodontitis e individuos sanos frente a proteínas de membrana externa de las cepas A. actinomycetemcomitans DSMZ8324, F. nucleatum DSM20482, P. gingivalis ATCC33277 y P. gingivalis W83.
Material y Métodos: Los cultivos de células planctónicas se obtuvieron con una suspensión bacteriana de 107 unidades formadoras de colonia (UFC)/mL en medio Brain Heart Infusion (BHI) modificado tras 24 horas de incubación. Después se procedió a la extracción de las proteínas bacterianas y a su cuantificación por el método Bradford. Posteriormente, se obtuvo el perfil proteico mediante electroforesis en geles de poliacrilamida, y se identificaron por espectrometría de masas. La respuesta de anticuerpos presentes en los sueros de los pacientes frente a las bacterias se estudió mediante la técnica de Western-blot. Resultados: La mayoría de las proteínas identificadas han mostrado ser de membrana externa. El estudio de los sueros de los distintos individuos mostró una respuesta variable de anticuerpos frente a las proteínas de las bacterias estudiadas, siendo de forma general mayor en los pacientes con periodontitis. Conclusiones: La optimización de un protocolo de extracción para el enriquecimiento en proteínas bacterianas de membrana externa ha quedado evidenciado. La respuesta inmunitaria de pacientes con periodontitis e individuos sanos frente a proteínas de membrana externa de A. actinomycetemcomitans, F. nucleatum y P. gingivalis ha sido variable y diferente entre los pacientes, no dependiendo siempre de la carga bacteriana que presenta el individuo.
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