TNFRp75-dependent immune regulation of alveolar macrophages and neutrophils during early Mycobacterium tuberculosis and Mycobacterium bovis BCG infection.

TNF signalling through TNFRp55 and TNFRp75, and receptor shedding is important for immune activation and regulation. TNFRp75 deficiency leads to improved control of Mycobacterium tuberculosis (M. tuberculosis) infection, but the effects of early innate immune events in this process are unclear. We investigated the role of TNFRp75 on cell activation and apoptosis of alveolar macrophages and neutrophils during M. tuberculosis and M. bovis BCG infection. We found increased microbicidal activity against M. tuberculosis occurred independently of IFNy and NO generation, and displayed an inverse correlation with alveolar macrophages (AMs) apoptosis. Both M. tuberculosis and M. bovis BCG induced higher expression of MHC-II in TNFRp75-/- AMs; however, M bovis BCG infection did not alter AM apoptosis in the absence of TNFRp75. Pulmonary concentrations of CCL2, CCL3 and IL-1ß were increased in TNFRp75-/- mice during M, bovis BCG infection, but had no effect on neutrophil responses. Thus, TNFRp75-dependent regulation of mycobacterial replication is virulence dependent and occurs independently of early alveolar macrophage apoptosis and neutrophil responses.

In vitro and in vivo toxicity evaluation of non-neuroleptic phenothiazines, antitubercular drug candidates.

The phenothiazine-derived antipsychotic drugs, such as chlorpromazine and thioridazine, are bactericidal against drug-sensitive and drug-resistant strains of Mycobacterium tuberculosis, but produce undesirable side effects at clinically relevant doses. We have previously modified four novel phenothiazines and maintained their antimycobacterial activity. This study evaluated the pharmacological and toxicity profiles of these novel non-neuroleptic phenothiazines, PTZ3, PTZ4, PTZ31 and PTZ32, for their metabolic stability, kinetic solubility and potential cytotoxic effects in vitro. To further support the safet use of these drug candidates, the in vivo pharmacological and toxicity profiles were assessed in C57BL/6 mice via single or repeated oral gavage. In acute toxicity studies, all four modified phenothiazines showed favourable safety in mice. When treated daily with 100 mg/kg of PTZ3 and PTZ4 for 2 weeks, mice displayed no signs of toxicity. Alternatively, treatment with PTZ31 resulted in 20% mortality with no toxicity evident in biochemical or histological analysis, while exposure to PTZ32 resulted in a 45% survival with increased serum concentrations of uric acid and alkaline phosphatase. The combined non-neuroleptic and antimycobacterial effects of the novel phenothiazines PTZ3, PTZ4, PTZ31 and PTZ32 demonstrated favourable pharmacological and toxicity profiles in this study, highlight the potential of these compounds as suitable anti-tuberculosis drug candidates.

Neurons are host cells for Mycobacterium tuberculosis.

Mycobacterium tuberculosis infection of the central nervous system is thought to be initiated once the bacilli have breached the blood brain barrier and are phagocytosed, primarily by microglial cells. In this study, the interactions of M. tuberculosis with neurons in vitro and in vivo were investigated. The data obtained demonstrate that neurons can act as host cells for M. tuberculosis. M. tuberculosis bacilli were internalized by murine neuronal cultured cells in a time-dependent manner after exposure, with superior uptake by HT22 cells compared to Neuro-2a cells (17.7% versus 9.8%). Internalization of M. tuberculosis bacilli by human SK-N-SH cultured neurons suggested the clinical relevance of the findings. Moreover, primary murine hippocampus-derived neuronal cultures could similarly internalize M. tuberculosis. Internalized M. tuberculosis bacilli represented a productive infection with retention of bacterial viability and replicative potential, increasing 2- to 4-fold within 48 h. M. tuberculosis bacillus infection of neurons was confirmed in vivo in the brains of C57BL/6 mice after intracerebral challenge. This study, therefore, demonstrates neurons as potential new target cells for M. tuberculosis within the central nervous system.

Synergistic Antimycobacterial Actions of Knowltonia vesicatoria (L.f) Sims.

Euclea natalensis A.DC., Knowltonia vesicatoria (L.f) Sims, and Pelargonium sidoides DC. are South African plants traditionally used to treat tuberculosis. Extracts from these plants were used in combination with isoniazid (INH) to investigate the possibility of synergy with respect to antimycobacterial activity. The ethanol extract of K. vesicatoria was subjected to fractionation to identify the active compounds. The activity of the Knowltonia extract remained superior to the fractions with a minimum inhibitory concentration (MIC) of 625.0 µg/mL against Mycobacterium smegmatis and an MIC of 50.00 µg/mL against M. tuberculosis. The K. vesicatoria extract was tested against two different drug-resistant strains of M. tuberculosis, which resulted in an MIC of 50.00 µg/mL on both strains. The combination of K. vesicatoria with INH exhibited the best synergistic antimycobacterial activity with a fractional inhibitory concentration index of 0.25 (a combined concentration of 6.28 µg/mL). A fifty percent inhibitory concentration of this combination against U937 cells was 121.0 µg/mL. Two compounds, stigmasta-5,23-dien-3-ol (1) and 5-(hydroxymethyl)furan-2(5H)-one (2), were isolated from K. vesicatoria as the first report of isolation for both compounds from this plant and the first report of antimycobacterial activity. Compound (1) was active against drug-sensitive M. tuberculosis with an MIC of 50.00 µg/mL.