The role of Toll-like Receptor 4 in the Modulation in Skeletal Muscle Metabolism
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Toll-like receptor 4 (TLR4) is a transmembrane receptor, which upon activation by lipopolysaccharide (LPS) from Gram-negative bacteria, plays an important role in the induction of the innate immune response. Our lab has previously demonstrated that activation of TLR4 in skeletal muscle results in the preferential oxidation of glucose for ATP production over that of fatty acids. Currently, the exact mechanism(s) for TLR4-induced modulation of metabolism are not known. The purpose of this project was to test the hypothesis that activation of TLR4 pathway causes increased ROS production, which contributes to deceased fatty acid oxidation and altered mitochondrial respiration in skeletal muscle. To this end, skeletal muscle cells were studied following acute and chronic treatments with LPS, and a mouse model with muscle-specific over expression of TLR4 (mTLR4) was studied under chow fed conditions and following 16 weeks of high fat feeding. Acute LPS treatment of C2C12 cells resulted in mitochondrial uncoupling as evidenced by higher levels of state IV respiration, reduced maximally simulated respiration, and a robust induction of uncoupling protein 3. These observations occurred in conjunction with increased pyruvate dehydrogenase activity. The LPS-induced changes in substrate preferences and maximally-stimulated mitochondrial respiration were prevented in the presence of the antioxidants, N-acetyl-L-cyteine (NAC) and catalase. Using isolated flexor digitorum brevis (FDB) muscle fibers from C57BL/6J mice, we showed that LPS treatment results in significant increases in ROS production that are evident at 15 min and still increasing at 45 min following the addition of LPS to incubation media. Hyperpolarization of mitochondrial membrane potential was also evident at 15 min post LPS treatment in FDB fibers. Fatty acid oxidation measured in skeletal muscle whole homogenates from the mTLR4 mice was significantly reduced compared to wild-type littermates on a standard chow diet. Following a 16 week high fat diet, the mTLR4, compared to wild-type mice, gained more weight and fat mass, were glucose intolerant, and displayed elevated production of mitochondrial-derived reactive oxygen species (ROS) from complex III. In conclusion, these data show that TLR4 activation elicits a change in mitochondrial substrate preference in that acetyl-CoA derived from pyruvate oxidation is the preferred substrate for the TCA cycle over that derived from Î²-oxidation of fatty acids. These data also lend strong support to the idea that the TLR4-mediated change in substrate preference is dependent on the production of ROS.
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