Morphological characterization and immunohistochemical detection of the proinflammatory cytokines IL-1β, IL-17A, and TNF-α in lung lesions associated with contagious bovine pleuropneumonia

Abstract

Contagious bovine pleuropneumonia (CBPP), a severe respiratory disease, is characterized by massive inflammation of the lung especially during the acute clinical stage of infection. Tissue samples from cattle, experimentally infected with Mycoplasma mycoides subsp. mycoides Afadé, were subjected to histopathological and immunohistochemical examination in order to provide insight into innate immune pathways that shape inflammatory host responses. Lung lesions were characterized by vasculitis, necrosis, and increased presence of macrophages and neutrophils, relative to uninfected animals. The presence of three cytokines associated with innate inflammatory immune responses, namely, IL-1β, IL-17A, and TNF-α, were qualitatively investigated in situ. Higher cytokine levels were detected in lung tissue samples from CBPP-affected cattle compared to samples derived from an uninfected control group. We therefore conclude that the cytokines TNF-α and IL-1β, which are prevalent in the acute phase of infections, play a role in the inflammatory response seen in the lung tissue in CBPP. IL-17A gets released by activated macrophages and attracts granulocytes that modulate the acute phase of the CBPP lesions.

Keywords: CBPP; Contagious bovine pleuropneumonia; Cytokines; Histopathology; Immunocytochemistry; Mycoplasma mycoides subsp. mycoides.

Fig. 1

a Lung—a shows severe, necrotizing bronchiolitis, with severe dense cellular, necrotic debris obstructing the bronchiolar lumen. The remaining bronchiolar wall shows loss of epithelial cells and severe infiltration of pulmonary and interstitial mixed inflammatory infiltrate with loss of pulmonary structure. Arrows indicate delineation of bronchiolar lining. b Lung—b shows intraalveolar proteinaceous, fibrinous precipitates (arrows) with loss of pulmonary architecture as well as thickened alveolar septa with mixed inflammatory, interstitial infiltrates within alveolar septa and interstitial space. c Lung—c shows a severe, necrotizing vasculitis of a pulmonary arteriole with loss of vascular wall (arrows indicate remnants of necrotic vascular wall) and severe, necrotic, cellular debris obliterating the vascular lumen. The perivascular interstitial space shows perivascular, mixed inflammatory cells and protein precipitates. d Lung—d shows perivascular pulmonary fibrosis extending into the interstitial space, with immature extracellular matrix and mixed inflammatory cells. Arrow indicates arterial media hypertrophy. e Lung—e shows an intrapulmonary arteriole with an adhesive, lumen-obstructing thrombus and media hypertrophy. Arrow indicates wall of arteriole. f Lung—f shows severe chronic, diffuse pleuritis with extensive subpleural fibrosis and neovascularization and loss of pulmonary architecture. Arrow indicates pleural surface. g Lung—g shows an interstitial inflammatory infiltrate composed of neutrophilic and eosinophilic granulocytes (arrows) within the interstitial space of the lung. h Pulmonary lymphnode—h shows a pulmonary lymph node with follicular hyperplasia and activated germinal centers (arrows). Size standards are displayed in the lower right corner of each picture: black represents 100 μm; grey represents 50 μm, and yellow represents 20 μm

Fig. 2

Representative qualitative results of the detection of the cytokines IL-1β, IL-17A, and TNF-α. The following polyclonal antibodies were used. a Anti- IL-1β: Lung: control animal; b Anti- IL-1β: Lung: acute CBPP lesion; c Anti- IL-17A: Lung: control animal; d Anti- IL-17A: Lung: acute CBPP lesion; e Anti- TNF-α: Lung: control animal; f Anti- TNF-α: Lung: acute CBPP lesion; g Anti- Mycoplasma: Lung: control animal; h Anti- Mycoplasma: Lung: acute CBPP lesion. Black bars in the lower right corner of images in a–f is the size standard representing 100 μm, black bars in images of g, h represent 20 μm. Arrows indicate positive immunoreactive signals (reddish-brown)