Expression and function of toll-like receptor 4 and inflammasomes in cardiac fibroblasts and myofibroblasts: IL-1β synthesis, secretion, and degradation.

UNLABELLED: Cardiac inflammation can be produced by pathogen-associated molecular patterns (PAMPs), from parasitic, bacterial or viral origin; or by danger-associated molecular patterns (DAMPs), released from dead cells after cardiac tissue damage, for example by cardiac infarction. Both, PAMPS and DAMPS activate TLR4 on resident immune cells and heart tissue cells, triggering an inflammatory process necessary to begin the wound healing process. Cardiac fibroblasts (CF) are the most abundant cells in the heart and are critical to wound healing, along with cardiac myofibroblasts (CMF), which are differentiated from CF through a TGF-β1-mediated process. While TLR4 and the inflammasome complex are known to play important roles in CF function, the effects of TGF-β1 on TLR4 and inflammasome expression and activity remain unknown. To elucidate this important point, we evaluated the effect of TGF-β1 on TLR4, and the inflammasome protein expression and activity through activation by LPS, mimicking a myocarditis condition by bacterial origin. We found that TGF-β1 increased TLR4 expression in CF and that the process was mediated by the TGFβRI and p38 signaling pathways. In both CF and CMF, LPS triggered ERK1/2, PI3K-Akt, and p65-NF-κB phosphorylation. All of these effects were blocked by TAK-242, a TLR4 signaling pathway inhibitor. LPS increased pro-IL-1β levels, which were dependent on the ERK1/2, PI3K-Akt, and NF-κB signaling pathways, and levels were higher in CF than CMF. NLRP3 and ASC levels were similar in CF and CMF, while pro-caspase-1 levels and caspase-1 activity were higher in CMF. LPS+ATP treatment induced inflammasome complex assembly and activation, triggering the release of IL-1β in both CMF and CF. Finally, the unsecreted pro-IL-1β in the CF was degraded by autophagy.

CONCLUSION: TGF-β1 increases TLR4 expression in CF. Despite different pro-IL-1β and caspase-1 activity levels in CF versus CMF, the two cell types secreted similar levels of IL-1β after LPS+ATP treatment. These findings suggest that both cell types are active participants in inflammation.