MRP2 inhibition mitigates pulmonary burden and bacteremia following lung challenge with S. pneumoniae. MRP2 inhibition mitigates pulmonary burden and bacteremia.

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MRP2 inhibition mitigates pulmonary burden and bacteremia following lung challenge with S. pneumoniae. MRP2 inhibition mitigates pulmonary burden and bacteremia following lung challenge with S. pneumoniae. Using an in vivo model of PMN migration, we examined the results of MRP2 inhibition. C57BL/6 mice were treated with either PBS or probenecid 3 h before and 3 h after intratracheal application of 2.5 × 105 S. pneumoniae. Four sets of mice were infected in this way. (A) Comparing the number of PMNs found in the lumen after isolating BAL fluid samples after 24 and 48 h postinfection. PMNs, identified as Ly6g-positive cells, were quantified by flow cytometry (n = 8). (B) Overall lung burden from mice sacrificed on day 2. Mice were sacrificed, lungs were excised and homogenized, and total bacterial burden was calculated using serial dilutions (n = 24 for each condition). The presence (+) or absence (−) of bacteremia is indicated as follows: +, mice had detected levels of bacteria in blood 48 h postinfection; −, no bacteria were detected in blood during tail vein bleeds. (C) Bacteremia, as measured by tail vein bleeds, from the cohorts in panel B (see Materials and Methods). Detected events of colony formation on day 1 (D1) and day 2 (D2) (n = 24) are shown. The broken line represents the limit of detection, and as such, values for mice without visible bacteremia were represented as 100 CFU/ml, just below this level of detection. In panels B and C, statistical significance was calculated using the Mann-Whitney test. (D) Survival experiment with the fourth set of mice. There were 16 mice in a group for each condition. Statistical significance was calculated using Mantel-Cox test and Gehan-Breslow-Wilcoxon test. Probenecid treatment consistently increased survival by approximately 30 to 40% during survival experiments and often delayed symptoms, such as lethargy. Mouse experiments were repeated, and similar results were observed in at least two different experiments. Andrew Zukauskas et al. mSphere 2018; doi:10.1128/mSphere.00303-18