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. 2025 Aug 19;152(7):486-488.
doi: 10.1161/CIRCULATIONAHA.124.073023. Epub 2025 Aug 18.

Reformulation of the Necroptosis Pathway in Reperfused Myocardial Infarction

Dongze Qin #  1   2   3 , Radheshyam Modanwal #  1   2   3 , Madeeha Ghazi #  1   2   3 , J Jose Corbalan #  1   2   3 , Xiaotong F Jia  1   2   3 , Joshua L Axelrod  1   2   3 , Roneldine Mesidor  4 , Hee Jae Jang  5 , Claudio Humeres  1   6   3 , Anis Hanna  1   6   3 , Fahimeh Varzideh  1   7   3 , Michelle D Tallquist  8 , Gaetano Santulli  1   7   3 , Qinghang Liu  9 , Lorrie A Kirshenbaum  10 , Hongmin Ni  11 , Timothy J Kamp  5 , Rebekah L Gundry  4 , Siddharth Balachandran  12 , Alexei Degterev  13 , John W Calvert  14 , Richard N Kitsis  1   2   3
Affiliations

Reformulation of the Necroptosis Pathway in Reperfused Myocardial Infarction

Dongze Qin et al. Circulation. .
No abstract available

Keywords: cell death; myocardial infarction; necroptosis.

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Conflict of interest statement

None.

Figures

Figure.
Figure.. Necroptosis during reperfused myocardial infarction is mediated by ZBP1 → RIPK3 → MLKL, rather than RIPK1 → RIPK3 → MLKL, signaling.
A-C. Immunoblots showing that RIPK1 is undetectable, or barely detectable, in adult murine and human cardiomyocytes. D, E. Absence of effect of cardiomyocyte-specific (Myh6-Mer-Cre-Mertg/_ ; Ripk1flox/flox) or fibroblast-specific (Tcf21-Mer-Cre-MerKI/+; Ripk1flox/flox) deletion of Ripk1 on infarct size in response to 45 min left coronary artery distribution ischemia/24 h reperfusion in vivo. Area at risk (AAR) and infarct assessed by Evans blue and tetrazolium chloride staining, respectively. F, G. Infarct size and RIPK3 and MLKL phosphorylation were not affected by homozygous germline knockin of kinase-dead Ripk1 allele (K45A). H. Necrostatin-1 (1.65 mg/kg into LV cavity immediately before reperfusion) cardioprotects to similar extents in control and cardiomyocyte-specific Ripk1 knockout mice. I, J. Germ line deletion of Ripk3 and Mlkl decreases infarct size. K, L. ZBP1 is transcriptionally induced in the heart following 45 min ischemia/4 h reperfusion. N=3 biological replicates per condition. M. ZBP1 levels increase in adult murine cardiomyocytes subjected to hypoxia (1h) and reoxygenation for the times indicated (AnaeroPak system). N=5–6 biological replicates per time point for wild type cardiomyocytes and 3 biological replicates for knockout cardiomyocytes, N. RIPK3 co-immunoprecipitates with ZBP1 in the heart. Mice homozygous for 3 x FLAG knockin into Zbp1 used as indicated. Example representative of 5 independent experiments. O, P. Germ line Zbp1 deletion reduces phosphorylation of RIPK3 and MLKL in response to MI/R. N=3 biological replicates per condition. Panel O is a phos-tag gel. Q. Zbp1 deletion reduces infarct size during MI/R. R. Zbp1 deletion reduces cell death in adult mouse cardiomyocytes subjected to 1 h hypoxia/4 h reoxygenation. N=4–5 biological replicates per condition. In vivo experiments employed 12–16-week age male mice. All mice are on a C57BL/6J background except Mlkl KO mice and their controls which were C57BL/6N. The surgeon was blinded to genotype both during the surgery and analysis of infarcts. Infarct size was assessed at the 24 h reperfusion time point in all experiments. Data from the 179/204 (87.7%) mice that exhibited the pre-specified AAR 40–65%, myocardial pallor with coronary occlusion and restoration of myocardial color upon suture release, and that survived to the sacrifice time point are reported. Signaling changes, which are manifested earlier, were assessed at the 4 h reperfusion time point in all experiments. Data displayed as mean ± SEM. Statistics: Panels D, E, F, I, J, Q – two-tailed Student’s t-test. Panels H, L, O, R – two-way ANOVA with Tukey’s test. *, **, ***, **** – P < 0.05, 0.01, 0.001, and 0.0001, respectively. Scale bar – 1 mm. RIPK1 – receptor interacting protein kinase 1. GAPDH – glyceraldehyde 3-phosphoate dehydrogenase. cTnT – cardiac troponin T. CM – cardiomyocyte. CF- cardiac fibroblast. AAR – area at risk. LV – left ventricle. INF – infarct. Nec-1 – necrostatin −1. RIPK3 – receptor interacting protein kinase 3. MLKL- mixed lineage kinase domain-like. ZBP1 – Z-nucleic acid binding protein 1. MI/R – myocardial infarction with reperfusion. H/R – hypoxia/reoxygenation. KO – knockout. K/I – knockin. All primary data, analytical methods, and materials will be made available to other researchers and can be obtained by contacting corresponding author RNK.
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