Differential cytokine gene expression in granulomas from lungs and lymph nodes of cattle experimentally infected with aerosolized Mycobacterium bovis

Authors: Palmer, Mitchell; Thacker, Tyler; Waters, Wade

Submitted to: PLOS ONE
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/16/2015
Publication Date: 12/17/2015
Citation: Palmer, M.V., Thacker, T.C., Waters, W.R. 2015. Differential cytokine gene expression in granulomas from lungs and lymph nodes of cattle experimentally infected with aerosolized Mycobacterium bovis. PLoS One. 10(12):e0142287.

Interpretive Summary: The causes of tuberculosis in humans and animals have been known for over a century. Since the discovery of tuberculosis scientists have worked to improve diagnostic tests to identify humans and animals with tuberculosis and produce vaccines to prevent tuberculosis. This requires a thorough understanding of the disease process. In humans and animals tuberculosis is characterized by the formation of granulomas; specific host responses (lesions) in infected organs, most commonly lymph nodes and lungs. To understand tuberculosis requires an understanding of the interaction of host and pathogen within the granuloma. We used cattle experimentally infected with Mycobacterium bovis to evaluate the inflammatory milieu inside granulomas of the lung and lymph nodes and showed that the inflammatory milieu within granulomas of lung and lymph node are very different, even though the granulomas are similar in microscopic appearance. We also showed that the inflammatory milieus within granulomas of two different lymph nodes associated with the lung are very different. This work demonstrates that although granulomas may look much alike, they can differ greatly based on where they are found; highlighting the need to select carefully those tissues being analyzed for immune studies of tuberculosis.

Technical Abstract: The hallmark lesion of tuberculosis in humans and animals is the granuloma. The granuloma represents a distinct host cellular immune response composed of epithelioid macrophages, lymphocytes, and multinucleated giant cells, often surrounding a caseous necrotic core. Within the granuloma, host interacts with pathogen and disease outcome is determined. Factors within the granulomas such as cytokines and chemokines drive cell recruitment, activity, function and ultimately the success or failure of infection control by the host. Hence, an understanding of the granuloma-level cytokine response is necessary to understand tuberculosis pathogenesis. In-situ cytokine expression patterns were measured using a novel in situ hybridization assay, known as RNAScope®, in granulomas of the lungs, tracheobronchial lymph nodes and caudal mediastinal lymph nodes of cattle experimentally infected with Mycobacterium bovis via aerosol exposure. In spite of microscopic morphological similarities, significant differences were seen between late stage granulomas of the lung compared to those of the tracheobronchial lymph nodes for IFN-gamma, IL10, IL-17A, IL-22 and TGF-beta, but not for TNF-alpha. Additionally, significant differences were noted between granulomas from two different thoracic lymph nodes that both receive afferent lymphatics from the lungs (i.e., tracheobronchial and caudal mediastinal lymph nodes) for IFN-gamma, TNF-alpha, IL-10, IL-17A and TGF-beta, but not for IL-22. These findings show that granuloma morphology alone is not a reliable indicator of granuloma function as granulomas of similar morphologies can have disparate cytokine expression patterns. Moreover, anatomically distinct lymph nodes (tracheobronchial vs caudal mediastinal) differ in cytokine expression patterns even when both receive afferent lymphatics from a lung containing tuberculoid granulomas. These findings show that selection of tissue and anatomic location are critical factors in assessing host immune response to M. bovis and should be considered carefully.