Abstract
Background:
Regulated forms of necrosis-like cell death (e.g., necroptosis) have been shown to contribute to cardiac ischemia/reperfusion (I/R) injury. However, pro-inflammatory necroptosis is unlikely to be involved during early reperfusion and little is known about the associated molecular changes. Thus, this study aimed to provide an in-depth protein screening with a particular focus on pro-pyroptotic and mitochondrial damage-related pathways.
Methods:
Langendorff-perfused rat hearts were subjected to 30-minute global ischemia followed by 10-minute reperfusion. Liquid chromatography coupled with mass spectrometry (LC-MS/MS) and immunoblotting techniques were used to study the complex cardiac proteome. In addition, calcium-induced mitochondrial swelling and lactate dehydrogenase (LDH) release were examined to assess mitochondrial stress and necrosis phenotype, respectively.
Results:
Approximately 160 proteins linked to cell death signaling, cellular metabolism, and post-translational modifications were significantly differentially expressed in I/R hearts compared to controls. Conventional proteins of pyroptosis, either of canonical or non-canonical signaling, were not affected during the short reperfusion. Notably, this type of I/R was associated with increased expression of p25 cleaved form of poly [ADP-ribose] polymerase 1 (PARP1 p25) and mature apoptosis-inducing factor (AIF), alongside nitrosative stress and mitochondrial swelling. Conversely, a receptor-interacting protein kinase 3 (RIP3) inhibitor (GSK′872, 250 nM) reversed mitochondrial swelling and plasma membrane rupture and mitigated the increase in the expression of PARP1 p25 and AIF.
Conclusions:
This study shows for the first time that necrosis-like injury during early I/R of the isolated heart is associated with mitochondrial events, rather than pro-inflammatory pyroptotic cell death. Furthermore, the inhibition of RIP3 mitigates this injury independent of targeting pro-inflammatory signaling.
Regulated forms of necrosis-like cell death (e.g., necroptosis) have been shown to contribute to cardiac ischemia/reperfusion (I/R) injury. However, pro-inflammatory necroptosis is unlikely to be involved during early reperfusion and little is known about the associated molecular changes. Thus, this study aimed to provide an in-depth protein screening with a particular focus on pro-pyroptotic and mitochondrial damage-related pathways.
Methods:
Langendorff-perfused rat hearts were subjected to 30-minute global ischemia followed by 10-minute reperfusion. Liquid chromatography coupled with mass spectrometry (LC-MS/MS) and immunoblotting techniques were used to study the complex cardiac proteome. In addition, calcium-induced mitochondrial swelling and lactate dehydrogenase (LDH) release were examined to assess mitochondrial stress and necrosis phenotype, respectively.
Results:
Approximately 160 proteins linked to cell death signaling, cellular metabolism, and post-translational modifications were significantly differentially expressed in I/R hearts compared to controls. Conventional proteins of pyroptosis, either of canonical or non-canonical signaling, were not affected during the short reperfusion. Notably, this type of I/R was associated with increased expression of p25 cleaved form of poly [ADP-ribose] polymerase 1 (PARP1 p25) and mature apoptosis-inducing factor (AIF), alongside nitrosative stress and mitochondrial swelling. Conversely, a receptor-interacting protein kinase 3 (RIP3) inhibitor (GSK′872, 250 nM) reversed mitochondrial swelling and plasma membrane rupture and mitigated the increase in the expression of PARP1 p25 and AIF.
Conclusions:
This study shows for the first time that necrosis-like injury during early I/R of the isolated heart is associated with mitochondrial events, rather than pro-inflammatory pyroptotic cell death. Furthermore, the inhibition of RIP3 mitigates this injury independent of targeting pro-inflammatory signaling.
| Original language | English |
|---|---|
| Article number | 27119 |
| Pages (from-to) | 27119 |
| Number of pages | 1 |
| Journal | Frontiers in Bioscience-Landmark |
| Volume | 30 |
| Issue number | 2 |
| DOIs | |
| Publication status | Published - 24 Jan 2025 |
Bibliographical note
Publisher Copyright:© 2025 The Author(s).