The Role Of Releasing Danger Signals In Atherosclerosis And Myocardial Infarction

Aim:The present study aimed to elucidate the mechanism by which bone marrow mesenchymal stem cells (BMSCs) differentiate into smooth muscle cells (SMCs) in atherosclerosis.Main methods:We isolated mouse BMSCs and incubated them in conditioned medium from plaque-derived SMCs (SMC-CM) and analyzed growth factors from media. BMSCs were treated with different media and harvested at continuous time points for investigating the ability to differentiate toward SMCs. Next, BMSCs of green fluorescence protein (GFP) mice were transplanted into apolipoprotein E(?)(apoE(?)) mice fed on western type diet for 12 weeks. In vivo efficacy of BMSCs was investigated.Key findings:After being cultured using SMC-CM, hepatocyte growth factor (HGF) was abundantly secreted into the medium by BMSCs with time. BMSCs had increased expression of HGF receptor c-met and SMC-specific markers while they also displayed SMC characteristic'hill and valley-like' appearance with an SMC ultra-structure including actin filaments and dense bodies. In vivo-grafted BMSCs aggravated atherosclerotic lesions and inflammation but ameliorated fibrosis in aorta while they displayed higher expression levels of c-met and early SMC-specific markers but not late-stage markers in aorta. They also demonstrated greater secretion of HGF in the aorta of apoE(?) mice. Furthermore, when BMSCs were treated with HGF blocking antibody, they lost the ability to differentiate to SMCs.Significance:HGF from local SMCs plays an important role for the differentiation of homing BMSCs. Innate immune response is the most common cause of myocardial inflammation after transient ischemia and may contribute to injury, yet the signaling mechanisms leading to such response are not well understood. We have previously demonstrated that transient myocardial ischemia activates interleukine-1 receptor-associated kinase-1 (IRAK-1), a kinase critical for interleukin-1 and Toll-like receptor (TLR) signaling. Here we tested the hypothesis that myocardial ischemia activates IRAK-1 via a mechanism that involves stress proteins such as heat shock proteins (HSPs) and TLRs. Coronary artery occlusion induced a rapid myocardial IRAK-1 activation within 30 min in wild type (WT), TLR2(?) or Trif(?) mice, but not in TLR4det or MyD88(?) mice. We found that HSP60 protein was significantly increased in serum or in perfusate of isolated heart after 30 min of myocardial ischemia. In vitro, recombinant HSP60 induced IRAK-1 activation in cells derived from WT, TLR2(?),or Trif(?) mice, but not from TLR4 or MyD88 mice. Both myocardial ischemia-and HSP60-induced IRAK-1 activation was abolished by HSP60 blocking antibody. Moreover, in the presence of serum, HSP60 treatment of cardiomyocytes led to marked activation of caspase-8 and-3, but not-9. Expression of dominant-negative mutant of Fas-associated death domain protein or a caspase-8 inhibitor completely blocked HSP60-induced caspase-8 activation. Finally, transgene expression of IRAK-1-WT, not its kinase-deficient mutant, induced caspase activation and apoptosis in cardiomyocytes cultured in serum-containing medium. Taken together, these data suggest that following transient ischemia, myocardium-released HSP60 induces IRAK-1 activation via TLR4→MyD88 signaling and that HSP60 elicits cardiomyocyte apoptosis by activating death-receptor pathway.