Fig 8. Specificity and efficacy of Oxa33.
(A) Rv and Rv::glmU tet-on cultures were grown in presence of 1 μg/ml ATc and different concentrations of Oxa33 or INH (as indicated). 0.02% resazurin dye was added on day 7. Pink colour indicates viable bacteria while blue colour indicates dead bacteria. ‘C’ denotes culture media control. The experiment was performed thrice. (B) Differentiated THP1 were infected either with Rv or Rv::glmU tet-on and grown in RPMI media containing 400 ng/ml ATc. 24 h post infection different concentrations (as indicated) of Oxa33 or vehicle control was supplemented to the media for the next 72 h followed by CFU enumeration. Experiment was performed in triplicates and the error bars represent s.e.m. (C) BALB/c mice (7 to 11 mice / group) were infected with Rv and the infection was allowed to be established for 28 day. Oxa33 (50 mg/kg, intraperitoneal, in 2.5% Tween 80) or INH (25 mg/kg, in drinking water with 5% sucrose) were administered to the mice for a subsequent period of 56 days. CFU in the lungs of mice were determined on 1, 28 and 84 days post infection. Day 1st and day 28th CFUs show successful implantation and establishment of infection, respectively. 56 days of Oxa33 or INH treatment partially reduces the mean bacillary load to 2.78 and 1.3 on log10 scale. ***p<0.0005 or 0.0006, two tailed non parametric t-test, error bars represent s.e.m. (D) While the overall pathology of lungs (upper panel) illustrates robust Mtb infection in untreated mice, relatively smaller and fewer lesions and granulomas were observed in Oxa33 or INH treated mice. Lower panel exhibit hematoxylin and eosin stained photomicrograph of lungs from Rv, INH treated and Oxa33 treated mice at 100x magnification. This histopathological analysis shows large or small granuloma with lymphocytes and foamy histiocytes in Rv infected mice or Oxa treated mice respectively G = Granuloma, BL = Bronchial Lumen, AS = Alveolar space. (E) Granuloma scores from lungs of infected and treated mice for 4 weeks and 12 weeks.