High and Low Levels of ABCB1 Expression Are Associated with Two Distinct Gene Signatures in Lung Tissue of Pulmonary TB Patients with High Inflammation Activity.

P-glycoprotein (encoded by the ABCB1 gene) has a dual role in regulating inflammation and reducing chemotherapy efficacy in various diseases, but there are few studies focused on pulmonary TB patients. In this study, our objective was to identify a list of genes that correlate with high and low levels of ABCB1 gene expression in the lungs of pulmonary TB patients with different activity of chronic granulomatous inflammation. We compared gene expression in two groups of samples (with moderate and high activity of tuberculomas) to identify their characteristic gene signatures. Gene expression levels were determined using quantitative PCR in samples of perifocal area of granulomas, which were obtained from 65 patients after surgical intervention. Subsequently, two distinct gene signatures associated with high inflammation activity were identified. The first signature demonstrated increased expression of HIF1a, TGM2, IL6, SOCS3, and STAT3, which correlated with high ABCB1 expression. The second signature was characterized by high expression of TNFa and CD163 and low expression of ABCB1. These results provide insight into various inflammatory mechanisms and association with P-gp gene expression in lung tissue of pulmonary TB patients and will be useful in the development of a host-directed therapy approach to improving the effectiveness of anti-TB treatment.

Application of Laser Scanning Confocal Microscopy for the Visualization of M. tuberculosis in Lung Tissue Samples with Weak Ziehl-Neelsen Staining.

One of the key requirements for the diagnosis of pulmonary tuberculosis is the identification of M. tuberculosis in tissue. In this paper, we present the advantages of specific fluorescent antibody labelling, combined with laser scanning confocal microscopy (LSCM), for the detection of M. tuberculosis in histological specimens of lung tissues. We demonstrate that the application of LSCM allows: (i) The automatic acquisition of images of the whole slice and, hence, the determination of regions for subsequent analysis; (ii) the acquisition of images of thick (20-40 µm) slices at high resolution; (iii) single bacteria identification; and (iv) 3D reconstruction, in order to obtain additional information about the distribution, size, and morphology of solitary M. tuberculosis; as well as their aggregates and colonies, in various regions of tuberculosis inflammation. LSCM allows for the discrimination of the non-specific fluorescence of bacteria-like particles and their aggregates presented in histological lung samples, from the specific fluorescence of labelled M. tuberculosis, using spectrum emission analysis. The applied method was effective in the identification of M. tuberculosis in lung histological samples with weak Ziehl-Neelsen staining. Altogether, combining immunofluorescent labelling with the application of LSCM visualization significantly increases the effectiveness of M. tuberculosis detection.