To determine changes in cellular activity within tissues due to viable or non-viable MAP and the introduction of NP-51 we preformed assays to measure host transcript expression for key inflammatory markers. Host immune cells may produce and store non-specific, pro-inflammatory cytokines in the event of infection and yield more specific cytokines as disease progresses. For these reasons, our evaluation of cytokine transcript concentrations was to determine their active production, find more post MAP infection. These results are highlighted in Figures 3 and 4, respectively. Figure 3 Serum

cytokine abundance relative to controls and associated with chronic MAP infection. Data for male and female animals and time points were combined for each experimental group (n = 24) for these results. Experimental groups analyzed were the following: control animals fed normal chow and uninfected (Control; C); animals fed normal chow and infected with non-viable MAP cells, (Killed-MAP; K-MAP); animals fed normal chow and infected with viable

MAP cells (Live-MAP; L-MAP); animals fed viable probiotics in chow and uninfected (Live NP-51; L-NP-51); animals fed viable probiotics in chow and infected with non- viable MAP cells (K-MAP + L-NP-51); animals fed viable probiotics in chow and infected with viable MAP cells (L-MAP + L-NP-51). Data analysis methods are further described in the data analysis section. Figure 4 Tissue cytokine transcript abundance relative to controls and associated with chronic MAP infection. Data for male and female animals, time points, GS-1101 in vivo and tissues (small/large intestine and liver) were combined for each experimental

group (n = 24). Experimental groups analyzed were the following: animal fed normal chow and uninfected (Control; C); animals fed normal chow and infected with non-viable MAP cells, (Killed-MAP; K-MAP); animals fed normal chow and infected with viable MAP cells (Live-MAP; L-MAP); animals fed viable probiotics in chow and uninfected (Live NP-51; L-NP-51); animals fed viable probiotics in chow and infected with non-viable MAP cells ( K-MAP + L-NP-51); PAK5 animals fed viable probiotics in chow and infected with viable MAP cells (L-MAP + L-NP-51). Data analysis methods are further described in the data analysis section. With viable MAP (L-MAP) infection, the immune response produced is characteristic of Th1 cell responses to intracellular pathogens with the production of IFN-Υ, IL-6, IL-12 (as described in Figure 3) [1, 2, 8]. In animals that were infected with viable MAP and fed viable probiotics (L- MAP + L-NP-51) – there is IFN-Υ production likely due to intracellular infection by MAP but this response is weaker compared to animals infected only with viable MAP, (see Figure 3). RXDX-101 ic50 Equally, IL-12 levels are elevated but with NP-51 consumption we again observe a decrease in IL-6 circulation and an increase in pro-inflammatory cytokine- TNF-α.