Project description:C3heB/FeJ mice were infected with M. tuberculosis to form necrotic granulomatous lesions. FFPE samples of infected lungs with granulomas were microdissected into three distinct regions, Caseum, foamy macrophage, and Cell. Proteins were extracted from microdissected samples, followed by LC-MS/MS.
Project description:We sought to develop and characterize a novel paucibacillary model in mice, which develop necrotic lung granulomas following infection with Mycobacterium tuberculosis. Paucibacillary infection was established, recapitulating the sterilizing activities of human LTBI regimens. TNF neutralization led to increased lung bacillary load, disrupted granuloma architecture with expanded necrotic foci and reduced tissue hypoxia, and accelerated animal mortality. TNF-neutralized mouse lungs and sera showed significant upregulation of IFN?, IL-1?, IL-6, IL-10, CCL2, CCL3, and matrix metalloproteinase genes Six weeks after aerosol-immunization with recombinant M. bovis BCG overexpressing the 30-kilodalton antigen, C3HeB/FeJ mice were aerosol-infected with M. tuberculosis H37Rv. Six weeks later, mice were treated with one of three standard regimens for latent TB infection (LTBI) or TNF-neutralizing antibody. Mouse lungs were analyzed by histology, positron emission tomography/computed tomography, whole-genome microarrays, and RT-PCR. Lungs and sera were studied by multiplex enzyme-linked immunosorbent assays
Project description:In-vivo Gene Signatures of Mycobacterium tuberculosis In C3HeB/FeJ Mice In this experiment we have compared Mtb transcriptomics in-vivo, using samples derived from chronically infected C3HeB/FeJ mice, which produce human like caseating lesions, unlike other murine species, to Mtb cultured in-vitro. Similarly, the genome-wide expression of MtbdeldosR, MtbdeldosS and MtbdeldosT mutants is also compared between lungs from C3HeB/FeJ mice and in-vitro culturing.
Project description:To gain holistic of the immune landscape of lesions following infection with Mycobacterium tuberculosis, we aerosol infected C3HeB/FeJ mice with 1-3 CFU of Mtb (SA161 strain). 35 days following infection, lungs were harvested and fixed, then embedded and frozed in OCT. 10um sections were obtained, classified as necrotic or non-necrotic by visual inspection and brightfield imaging, then stained with markers delineating lesions, and subsequently processed and sampled using the nanostring GeoMX platform. We then compared the transcriptional environment of necrotic vs non-necrotic lesions
Project description:Comparison of kidney and liver samples from wildtype C3HeB/FeJ mice. 16 dual-color DNA-chip hybridizations of cDNAs from three age-matched male mice were made. For each individual mouse 6 or 4 replicate hybridizations were done. Keywords = liver Keywords = kidney Keywords = C3HeB/FeJ Keywords: other
Project description:Granulomas function in humans during tuberculosis by focusing production of host antimicrobial factors against the causative bacterial agent Mycobacterium tuberculosis to contain infection. We show that mice unable to produce nitric oxide –itself an important antimicrobial molecule- demonstrate functional granulomas in the lung able to control infection after dermal infection. Disease in the lung was activated by administration of neutralising antibody against either TNF-α, which disrupted granuloma integrity, or INF-γ, which resulted in development of caseous necrosis within granulomas reminiscent of active human tuberculosis. In the latter case, the serpin protease inhibitor serpinb3a and its target protease, cathepsin G are highly expressed in cells local to necrotic regions in granulomas and serpinb3a induces necrosis of infected macrophages independently of cathepsin G binding. Therefore a single host protein is capable of inducing necrosis and bacterial growth during intracellular infection.
Project description:Neutrophils are accumulated in mouse lungs following Mycobacterium tuberculosis infections. Increased influx of neutrophils is associated with increased susceptibility to disease, but the underlying mechanisms of neutrophil mediated Tuberculosis disease pathogenesis is poorly understood. To understand the transcriptomic differences between resistant (C57 BL/6) and susceptible (C3HeB/FeJ) mouse lung neutrophils, we sorted them at Day 29 post infection and analyzed them by bulk mRNA sequencing.
