An in vitro human PBMC-based granuloma model as a drug screening, cellular function, and HIV/M.tb co-infection platform.
A histologic hallmark of tuberculosis (TB) is the granuloma, a unique multicellular tissue environment that both controls M. tuberculosis (M.tb) and fosters its persistence in macrophages leading to enhanced drug resistance and a propensity for reactivation. Granuloma structures prevent access to some antibiotics, facilitating bacterial survival. Most first-line anti-TB drugs have not shown a correlation between drug concentration in plasma and drug penetration in M.tb-induced lung granulomas, leading to suboptimal dosage. We have developed a human PBMC-based in vitro granuloma model containing virulent M.tb (mBio 6(1):e02537-14, 2015) designed for pathogenesis studies and translational applications, such as drug testing. We hypothesize this model will provide a screening tool that more accurately predicts drug efficacy in people. We generated granulomas with a luciferase reporter drug susceptible M.tb strain and two clinical drug resistant strains (isoniazid+ethambutol and rifampicin+streptomycin) using PBMCs from LTBI individuals. Antibiotics were then added at a range of concentrations. We compared our results with existing drug screening models and results to this point suggest that bacterial inhibition is achieved at higher antibiotic concentrations, when compared to in vitro MIC data or in vivo mouse models. Interestingly, bacterial inhibition activity of antibiotics with known low or no activity inside granuloma caseum was dose dependent. In other studies, we are evaluating the kinetics of granuloma formation using quantitative live cell microscopy and the impact of adding HIV-infected CD4 T cells. In summary, our human PBMC-based granuloma model shows promise as a novel and viable drug screening tool that recapitulates the human immune response which sequesters M.tb into granulomatous structures. We can also rapidly quantify granuloma formation and stability in an unbiased manner and elucidate the dynamics of granuloma formation and evolution, and the impact of HIV co-infection. NIAID R21 5R21AI145539-02.
Guerrero-Arguero I1, Renshaw C1, Arnett E1, Schlesinger S. L1
1Texas Biomedical Research Institute, San Antonio, TX