The Chemically-Modified Tetracycline COL-3 and Its Parent Compound Doxycycline Prevent Microglial Inflammatory Responses by Reducing Glucose-Mediated Oxidative Stress

We used mouse microglial cells in culture activated by lipopolysaccharide (LPS) or alpha-synuclein amyloid aggregates (alpha Sa) to study the anti-inflammatory effects of COL-3, a tetracycline derivative without antimicrobial activity. Under LPS or alpha Sa stimulation, COL-3 (10, 20 mu M) efficiently repressed the induction of the microglial activation marker protein Iba-1 and the stimulated-release of the pro-inflammatory cytokine TNF-alpha. COL-3 ' s inhibitory effects on TNF-alpha were reproduced by the tetracycline antibiotic doxycycline (DOX; 50 mu M), the glucocorticoid dexamethasone, and apocynin (APO), an inhibitor of the superoxide-producing enzyme NADPH oxidase. This last observation suggested that COL-3 and DOX might also operate themselves by restraining oxidative stress-mediated signaling events. Quantitative measurement of intracellular reactive oxygen species (ROS) levels revealed that COL-3 and DOX were indeed as effective as APO in reducing oxidative stress and TNF-alpha release in activated microglia. ROS inhibition with COL-3 or DOX occurred together with a reduction of microglial glucose accumulation and NADPH synthesis. This suggested that COL-3 and DOX might reduce microglial oxidative burst activity by limiting the glucose-dependent synthesis of NADPH, the requisite substrate for NADPH oxidase. Coherent with this possibility, the glycolysis inhibitor 2-deoxy-D-glucose reproduced the immunosuppressive action of COL-3 and DOX in activated microglia. Overall, we propose that COL-3 and its parent compound DOX exert anti-inflammatory effects in microglial cells by inhibiting glucose-dependent ROS production. These effects might be strengthened by the intrinsic antioxidant properties of DOX and COL-3 in a self-reinforcing manner.

Automated summary

The study showed that COL-3 and its parent compound DOX exert anti-inflammatory effects in microglial cells by inhibiting glucose-dependent ROS production. These effects were strengthened by the intrinsic antioxidant properties of DOX and COL-3 in a self-reinforcing manner, which also reduced microglial oxidative burst activity and TNF-alpha release in activated microglia.