Abstract 238: Toll-Like Receptor 4-Deficiency Improves Ischemic Cardiomyocytes Contractile Dysfunction via Augmentation of AMPK Activation

Abstract

Myocardial ischemia/reperfusion injury involves a robust inflammatory response especially activation of toll-like receptor 4 (TLR4), a proximal signaling receptor in innate immune responses to lipopolysaccharide of Gram-negative pathogens. TLR4 is expressed in the heart and vasculature, the deficiency of which was shown to protect the heart against ischemia/reperfusion injury. However, the molecular mechanisms behind TLR4 deficiency-induced cardioprotection against ischemic damage remain unclear. The AMP-activated protein kinase (AMPK), an intracellular energy gauge, plays an important role in limiting cardiac damage following by ischemia, thus we hypothesized that AMPK maybe a mediator of TLR4-deficiency cardioprotection against ischemic injury. To test this hypothesis, In vivo regional ischemia was induced by occlusion of the left anterior descending (LAD) coronary artery in wild type (WT) C3H/HeN and TLR4-deficient C3H/HeJ mice, which carry a natural mutation of TLR4. Immunoblotting analysis was performed to assess the activation of AMPK and TLR4 downstream signal, the mitogen-activated protein kinase (MAPK). The endoplasmic reticulum (ER) chaperons, Grp78/BiP and Gadd153/CHOP, and ER integral membrane protein IRE1α were also monitored. The results demonstrated that C3H/HeJ hearts had impaired ischemic MAPK and AMPK activation compared to WT C3H/HeN hearts, i.e. activation of p38 MAPK and JNK was blunted, while ERK and AMPK signaling pathways were augmented during ischemia in C3H/HeJ hearts. Intriguingly, ischemia-stimulated ER stress was higher in WT C3H/HeN hearts than that in C3H/HeJ as demonstrated by up-regulation of Grp78/BiP, Gadd153/CHOP and IRE1α. Caspase-3 activity and TUNEL staining results showed that WT C3H/HeN hearts have more damages than those of C3H/HeJ hearts. Moreover, isolated cardiomyocytes from C3H/HeJ hearts showed resistance to ischemia-induced contractile dysfunction compared to WT C3H/HeN hearts. These findings suggest that the cardioprotective effects against ischemic injury of TLR4-deficient hearts may mediated through augmentation of AMPK and ERK while alleviation of p38 MAPK and JNK inflammatory signaling pathways. This research has received full or partial funding support from the American Heart Association, AHA National Center.