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Figure 7 | Journal of Neuroinflammation

Figure 7

From: Reactive oxygen species and p47phox activation are essential for the Mycobacterium tuberculosis-induced pro-inflammatory response in murine microglia

Figure 7

Neither astrocytes nor indirect stimuli such as IL-1β adversely affected the s-Mtb-induced ROS release and cytokine production by primary mixed glial cells. A) ROS generation in astrocyte-enriched cultures. Astrocyte-enriched cultures were incubated with DHE (for superoxide detection) in the presence or absence of 1% s-Mtb or 100 ng/ml LPS for 30 min. The live cells were washed with serum-free medium and imaged using confocal microscopy. Images are representative of three independent experiments. B) Cytokine production in astrocyte-enriched cultures. Astrocyte-enriched cultures were stimulated with 1% s-Mtb for the indicated times (0- 48 h). The supernatant was analyzed for cytokine production using ELISA for TNF-α and IL-6. Values are the mean ± SD of triplicate samples. C) The effects of IL-1β on superoxide production. Primary mixed glial cells were stimulated with 1% s-Mtb or 100 ng/ml LPS in the presence of a neutralizing monoclonal antibody to IL-1β (1 μg/ml) or isotype control (1 μg/ml). After a 1-h incubation at 37°C, the cells were incubated with DHE (for superoxide detection) for 30 min. The live cells were washed with serum-free medium and imaged using confocal microscopy. The images are representative of three independent experiments. D) The effects of IL-1β on cytokine production. Primary mixed glial cells were pre-incubated with a neutralizing monoclonal antibody to IL-1β (1 μg/ml) or isotype control (1 μg/ml) in the presence or absence of NAC (20 mM), DPI (20 μM), rotenone (10 μM), allopurinol (100 μM), or 0.1% DMSO control for 30 min. The cells were then stimulated with 1% s-Mtb for 18 h. The supernatant was analyzed for cytokine production using ELISA for TNF-α and IL-6. Values are the mean ± SD of triplicate samples.

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