Tumor Necrosis Factor and Its Receptors Are Crucial to Control Mycobacterium bovis Bacillus Calmette-Guerin Pleural Infection in a Murine Model

Abstract

Tumor necrosis factor (TNF) is crucial to control Mycobacterium tuberculosis infection, which remains a leading cause of morbidity and mortality worldwide. TNF blockade compromises host immunity and may cause reactivation of latent infection, resulting in overt pulmonary, pleural, and extrapulmonary tuberculosis. Herein, we investigate the roles of TNF and TNF receptors in the control of Mycobacterium bovis bacillus Calmette-Guerin (BCG) pleural infection in a murine model. As controls, wild-type mice and those with a defective CCR5, a receptor that is crucial for control of viral infection but not for tuberculosis, were used. BCG-induced pleural infection was uncontrolled and progressive in absence of TNF or TNF receptor 1 (TNFR1)/TNFR2 (TNFR1R2) with increased inflammatory cell recruitment and bacterial load in the pleural cavity, and heightened levels of pleural and serum proinflammatory cytokines and chemokines, compared to wild-type control mice. The visceral pleura was thickened with chronic inflammation, which was prominent in TNF(-/-) and TNFR1R2(-/-) mice. The parietal pleural of TNF(-/-) and TNFR1R2(-/-) mice exhibited abundant inflammatory nodules containing mycobacteria, and these mice developed nonresolving inflammation and succumbed from disseminated BCG infection. By contrast, CCR5(-/-) mice survived and controlled pleural BCG infection as wild-type control mice. In conclusion, BCG-induced pleurisy was uncontrolled in the absence of TNF or TNF receptors with exacerbated inflammatory response, impaired bacterial clearance, and defective mesothelium repair, suggesting a critical role of TNF to control mycobacterial pleurisy.

Transmembrane Tumor Necrosis Factor Controls Myeloid-Derived Suppressor Cell Activity via TNF Receptor 2 and Protects from Excessive Inflammation during BCG-Induced Pleurisy

“…We have previously reported that a mouse model of pleural infection shows that TNF or its receptors are crucial to control M. bovis BCG-induced pleurisy ( 33 ). To further evaluate whether transmembrane TNF (tmTNF) or soluble-TNF form is required for protection during pleural mycobacterial infection, we analyzed mice that express a mutated form of TNF (tmTNF Δ1–9,K11E or tmTNF KI) that cannot be cleaved by TACE and do not produce solTNF ( 29 ).…”

“…Pleural BCG infection in TNF KO, but not tmTNF KI mice resulted in expansion of multinucleated giant cells containing numerous vacuoles and many bacilli (Figure 1 C). TNF KO cells were previously shown to be deficient in iNOS expression and unable to eliminate bacteria which led to a miliary TB ( 33 ). Accumulated cells were mainly myeloid CD11b + cells and the total number was not affected by the absence of TNF (Figure 1 D).…”

“…To examine if TNF signaling was also important for MDSC activity during acute BCG infection, we used sorted pleural MDSC from BCG-infected TNFR1/TNFR2 KO mice. These mice were shown to be highly sensitive to both systemic and pleural BCG infection ( 31 , 33 ). Both MO- and PMN-MDSC displayed complete absence of suppressive activity on CD4 T cells and even an enhancement of the proliferation of CD4 T cells co-cultured with MO-MDSC was observed (Figure S5 in Supplementary Material).…”

“…BCG-induced pleurisy infection was generated by intrapleural cavity injection of 10 6 CFU of M. bovis . BCG in 100 µL of saline as previously reported ( 33 ). Mice were monitored twice a week for body weight and sacrificed at day 14 post-infection or followed for survival studies.…”

“…Thoracic cavities from naïve and infected mice were washed twice with 1 mL of 2 mmol/L EDTA-phosphate-buffered saline (PBS), samples were centrifuged, and supernatants containing pleural fluid were frozen at −80°C for cytokine evaluation. Pleural cells were suspended in PBS-1% bovine serum albumin, counted, and used for the different techniques such as Flow cytometry, enrichment of MDSC, and cytospin followed by May-Grünwald-Giemsa and Ziehl-Neelsen (ZN) staining as reported ( 33 ).…”