Different signalling in infarcted and non‐infarcted areas of rat failing hearts: A role of necroptosis and inflammation

Abstract

Necroptosis has been recognized in heart failure (HF). In this study, we investigated detailed necroptotic signalling in infarcted and non‐infarcted areas separately and its mechanistic link with main features of HF. Post‐infarction HF in rats was induced by left coronary occlusion (60 minutes) followed by 42‐day reperfusion. Heart function was assessed echocardiographically. Molecular signalling and proposed mechanisms (oxidative stress, collagen deposition and inflammation) were investigated in whole hearts and in subcellular fractions when appropriate. In post‐infarction failing hearts, TNF and pSer229‐RIP3 levels were comparably increased in both infarcted and non‐infarcted areas. Its cytotoxic downstream molecule p‐MLKL, indicating necroptosis execution, was detected in infarcted area. In non‐infarcted area, despite increased pSer229‐RIP3, p‐MLKL was present in neither whole cells nor the cell membrane known to be associated with necroptosis execution. Likewise, increased membrane lipoperoxidation and NOX2 levels unlikely promoted pro‐necroptotic environment in non‐infarcted area. Collagen deposition and the inflammatory csp‐1‐IL‐1β axis were active in both areas of failing hearts, while being more pronounced in infarcted tissue. Although apoptotic proteins were differently expressed in infarcted and non‐infarcted tissue, apoptosis was found to play an insignificant role. p‐MLKL‐driven necroptosis and inflammation while inflammation only (without necroptotic cell death) seem to underlie fibrotic healing and progressive injury in infarcted and non‐infarcted areas of failing hearts, respectively. Upregulation of pSer229‐RIP3 in both HF areas suggests that this kinase, associated with both necroptosis and inflammation, is likely to play a dual role in HF progression.

Programmed Cell Death in the Left and Right Ventricle of the Late Phase of Post-Infarction Heart Failure

“…In spite of this experimental limitation, considering the decreased pSer229-RIP3 levels in the RV, it can be suggested, that despite MLKL upregulation, the post-MI pro-necroptotic environment in the LV spared the neighboring RV, at least in this stage of HF. Such an observation is in accordance with recent knowledge about necroptosis regulation, where particular changes in pSer229-RIP3 corresponded with p-MLKL, thereby indicating the presence or absence of necroptosis [ 7 , 8 , 13 , 15 ].…”

“…In our previous study, we have shown the activation of canonical necroptosis signaling being implicated in both infarcted and non-infarcted LV areas of failing rat hearts, while its execution mechanisms were restricted to the former area only. Such activation of necroptotic signaling has been suggested to be associated with myocardial fibrosis, ventricular remodeling, inflammation and severe contractile dysfunction [ 8 ]. Likewise, studies employing explanted human failing hearts of various etiologies have shown the participation of necroptosis in HF-associated myocardial deterioration [ 4 , 7 ].…”

“…Although we have found no changes in total RIP3 expression among the groups, its Ser229 phosphorylation, a specific pro-necroptotic phosphorylation, has been increased in the LV, but not in the RV of failing hearts. Because of methodological issues, as addressed in our most recent study [ 8 ], as well as discussed elsewhere [ 19 ], we assessed only total levels of MLKL instead of p-MLKL, a downstream target of pSer229-RIP3. Expression of MLKL in the LV was not affected by HF, but statistical analysis revealed significantly higher expression of MLKL in the RV of failing hearts.…”

“…Understanding the mechanisms of cell death in heart failure (HF) after myocardial infarction (MI) is of great clinical importance, since the extent of cellular loss significantly determines the degree of cardiac injury, contractile dysfunction, adverse remodeling and finally patient prognosis [ 1 , 2 , 3 ]. Therefore, an enormous effort has been undertaken to elucidate the role of certain cell death modalities in HF, and current knowledge on autophagy [ 4 , 5 , 6 ] and necroptosis [ 4 , 7 , 8 , 9 , 10 ], commonly occurring in post-infarction HF [ 4 , 5 , 7 , 8 , 9 , 10 ], has been advanced.…”

“…Besides the investigation of the left ventricle (LV) of failing hearts, which is primarily affected due to MI, we also examined the right ventricle (RV) because its functional impairment is a common clinical outcome of LV dysfunction. In addition, because apoptosis has been indicated as an important reason of cardiomyocyte death in HF [ 36 ], although recent data seem to undermine its significance [ 7 , 8 , 30 , 37 ], we analyzed some markers of this caspase-dependent cell death pathway as well.…”