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Pharmacological inhibition of TLR4 ameliorates muscle and liver ceramide content after disuse in previously physically active mice.

American journal of physiology. Regulatory, integrative and comparative physiology (2020-01-30)
Alec I McKenzie, Paul T Reidy, Daniel S Nelson, Jade L Mulvey, Nikol M Yonemura, Jonathan J Petrocelli, Ziad S Mahmassani, Trevor S Tippetts, Scott A Summers, Katsuhiko Funai, Micah J Drummond
ABSTRACT

Toll-like receptor 4 (TLR4) is a proposed mediator of ceramide accumulation, muscle atrophy, and insulin resistance in skeletal muscle. It is currently unknown whether pharmacological inhibition of TLR4, using the TLR4-specific inhibitor TAK-242 during muscle disuse, is able to prevent changes in intracellular ceramide species and consequently preserve muscle size and insulin sensitivity in physically active mice. To address this question, we subjected running wheel-conditioned C57BL/6 male mice (13 wk old; ∼10/group) to 7 days of hindlimb suspension (HS), 7 days of continued wheel running (WR), or daily injections of TAK-242 during HS (HS + TAK242) for 7 days. We measured hindlimb muscle morphology, intramuscular and liver ceramide content, HOMA-IR, mRNA proxies of ceramide turnover and lipid trafficking, and muscle fatty acid and glycerolipid content. As a result, soleus and liver ceramide abundance was greater (P < 0.05) in HS vs. WR but was reduced with TLR4 inhibition (HS + TAK-242 vs. HS). Muscle mass declined (P < 0.01) with HS (vs. WR), but TLR4 inhibition did not prevent this loss (soleus: P = 0.08; HS vs. HS + TAK-242). HOMA-IR was impaired (P < 0.01) in HS versus WR mice, but only fasting blood glucose was reduced with TLR4 inhibition (HS + TAK-242 vs HS, P < 0.05). Robust decreases in muscle Spt2 and Cd36 mRNA and muscle lipidomic trafficking may partially explain reductions in ceramides with TLR4 inhibition. In conclusion, pharmacological TLR4 inhibition in wheel-conditioned mice prevented ceramide accumulation during the early phase of hindlimb suspension (7 days) but had little effect on muscle size and insulin sensitivity.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
DL-Glyceraldehyde 3-phosphate solution, 45-55 mg/mL in H2O
Sigma-Aldrich
Anti-Laminin antibody produced in rabbit, 0.5 mg/mL, affinity isolated antibody, buffered aqueous solution
Sigma-Aldrich
TLR4 Inhibitor, TAK-242 - CAS 243984-11-4 - Calbiochem, TAK-242; CAS 243984-11-4; is a cell-permeable compound that selectively binds to Cys747 of TLR4 and selectively disrupts its interaction with adaptor molecules TIRAP and TRAM.