Sunil Patel, Thomas Francis, Raghini Rajaram, Rhodri Handslip, Sharon Mumby, Danielle E. Bear, Rahul Padhke, Nicholas Hart, Hugh Montgomery, Masao Takata, Stephen D.R. Harridge, Brijesh V. Patel, Zudin Puthucheary
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Samples were available from ICU admission (T1) and between day 7–10 post admission (T2). Skeletal muscle was stratified by a histopathologist in the original study as necrotic (NEC, <i>N</i> = 14) or non-necrotic (NONEC, <i>N</i> = 14) using haematoxylin and eosin staining. We used phosphorylated mixed-lineage kinase domain-like (pMLKL) protein (a key terminal effector protein) and receptor-interacting protein kinase 3 (RIPK3) as markers of necroptosis activation using Western blotting and immunohistochemistry.</p>\n </section>\n \n <section>\n \n <h3> Results</h3>\n \n <p>We show that pMLKL expression is significantly higher in the NEC group [NEC: T2:T1 expression; 9.1 (IQR 3.9–22.3) vs. NONEC: T2:T1 expression; 0.9 (IQR 0.6–1.1), <i>P</i> = 0.003]. We then confirm this upregulation and describe co-localization with receptor interacting protein kinase 3 (RIPK3) in skeletal muscle using immunohistochemistry. We show that both RIPK3 and pMLKL are present within intact myofibres at the intermediate timepoint day 3 without cellular infiltrate. At T2, pMLKL is also present in the interstitial space where there is infiltrate of CD68 positive immune cells. The observed necroptosis may originate from both internal and infiltrating sources. These findings were absent in samples from patients who did not exhibit histopathological features of necrosis.</p>\n </section>\n \n <section>\n \n <h3> Conclusions</h3>\n \n <p>We show that necroptosis machinery, RIPK3 and pMLKL, are associated with conventional histopathological features of myonecrosis in a critically ill cohort.</p>\n </section>\n </div>","PeriodicalId":73544,"journal":{"name":"JCSM rapid communications","volume":"6 2","pages":"111-121"},"PeriodicalIF":0.0000,"publicationDate":"2023-12-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/rco2.83","citationCount":"0","resultStr":"{\"title\":\"Programmed myofibre necrosis in critical illness acquired muscle wasting\",\"authors\":\"Sunil Patel, Thomas Francis, Raghini Rajaram, Rhodri Handslip, Sharon Mumby, Danielle E. Bear, Rahul Padhke, Nicholas Hart, Hugh Montgomery, Masao Takata, Stephen D.R. Harridge, Brijesh V. 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引用次数: 0
摘要
急性骨骼肌萎缩在危重疾病期间是常见的，并引起显著的发病率和功能限制。肌纤维坏死是一种主要的组织学发现，但通常被认为是肌肉炎症的非程序性副产品。本研究旨在评估是否一种形式的程序性坏死，坏死性上睑下垂，骨骼肌在危重疾病期间被激活。来自MUSCLE‐UK研究(ClinicalTrials.gov: NCT01106300)的28例患者的血清和骨骼肌活检样本被确定。样本可从ICU入院(T1)和入院后7-10天(T2)获得。在最初的研究中，组织病理学家使用血红素和伊红染色将骨骼肌分层为坏死(NEC, N = 14)或非坏死(NONEC, N = 14)。我们使用磷酸化的混合谱系激酶结构域样蛋白(pMLKL)(一种关键的末端效应蛋白)和受体相互作用蛋白激酶3 (RIPK3)作为necroptosis激活的标记物，使用Western blotting和免疫组织化学。我们发现pMLKL的表达在NEC组中显著升高[NEC: T2:T1表达;9.1 (IQR 3.9-22.3) vs. NONEC: T2:T1表达;0.9 (iqr 0.6 ~ 1.1)， p = 0.003]。然后，我们用免疫组织化学方法证实了这种上调，并描述了骨骼肌中受体相互作用蛋白激酶3 (RIPK3)的共定位。我们发现RIPK3和pMLKL在没有细胞浸润的情况下，在第3天的中间时间点存在于完整的肌纤维中。T2时，pMLKL也存在于CD68阳性免疫细胞浸润的间质间隙。观察到的坏死性下垂可能来源于内部和浸润性来源。这些发现在没有表现出坏死组织病理特征的患者样本中是不存在的。我们发现坏死坏死机制RIPK3和pMLKL与危重患者群体中肌坏死的常规组织病理学特征相关。
Programmed myofibre necrosis in critical illness acquired muscle wasting
Background
Acute skeletal muscle wasting during critical illness is common and causes significant morbidity and functional limitation. Myofibre necrosis is a major histological finding but is often considered an unprogrammed by-product of muscle inflammation. This study sought to evaluate if a form of programmed necrosis, necroptosis, is activated in skeletal muscle during critical illness.
Methods
A cohort of 28 patients from the MUSCLE-UK study (ClinicalTrials.gov: NCT01106300) with serum and skeletal muscle biopsy samples were identified. Samples were available from ICU admission (T1) and between day 7–10 post admission (T2). Skeletal muscle was stratified by a histopathologist in the original study as necrotic (NEC, N = 14) or non-necrotic (NONEC, N = 14) using haematoxylin and eosin staining. We used phosphorylated mixed-lineage kinase domain-like (pMLKL) protein (a key terminal effector protein) and receptor-interacting protein kinase 3 (RIPK3) as markers of necroptosis activation using Western blotting and immunohistochemistry.
Results
We show that pMLKL expression is significantly higher in the NEC group [NEC: T2:T1 expression; 9.1 (IQR 3.9–22.3) vs. NONEC: T2:T1 expression; 0.9 (IQR 0.6–1.1), P = 0.003]. We then confirm this upregulation and describe co-localization with receptor interacting protein kinase 3 (RIPK3) in skeletal muscle using immunohistochemistry. We show that both RIPK3 and pMLKL are present within intact myofibres at the intermediate timepoint day 3 without cellular infiltrate. At T2, pMLKL is also present in the interstitial space where there is infiltrate of CD68 positive immune cells. The observed necroptosis may originate from both internal and infiltrating sources. These findings were absent in samples from patients who did not exhibit histopathological features of necrosis.
Conclusions
We show that necroptosis machinery, RIPK3 and pMLKL, are associated with conventional histopathological features of myonecrosis in a critically ill cohort.
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