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Mitochondrial dynamics and mitophagy are constitutive and complex systems that ensure a healthy mitochondrial network through the segregation and subsequent degradation of damaged mitochondria. Disruption of these systems can lead to mitochondrial dysfunction and has been established as a central mechanism of ischemia/reperfusion (I/R) injury. Emerging evidence suggests that mitochondrial dynamics and mitophagy are integrated systems; however, the role of this relationship in the context of I/R injury remains unclear. To investigate this concept, we utilized primary cortical neurons isolated from the novel dual-reporter mitochondrial quality control knockin mice (C57BL/6-Gt(ROSA)26Sortm1(CAG-mCherry/GFP)Ganl/J) with conditional knockout (KO) of Drp1 to investigate changes in mitochondrial dynamics and mitophagic flux during in vitro I/R injury. Mitochondrial dynamics was quantitatively measured in an unbiased manner using a machine learning mitochondrial morphology classification system, which consisted of four different classifications: network, unbranched, swollen, and punctate. Evaluation of mitochondrial morphology and mitophagic flux in primary neurons exposed to oxygen-glucose deprivation (OGD) and reoxygenation (OGD/R) revealed extensive mitochondrial fragmentation and swelling, together with a significant upregulation in mitophagic flux. Furthermore, the primary morphology of mitochondria undergoing mitophagy was classified as punctate. Colocalization using immunofluorescence as well as western blot analysis revealed that the PINK1/Parkin pathway of mitophagy was activated following OGD/R. Conditional KO of Drp1 prevented mitochondrial fragmentation and swelling following OGD/R but did not alter mitophagic flux. These data provide novel evidence that Drp1 plays a causal role in the progression of I/R injury, but mitophagy does not require Drp1-mediated mitochondrial fission.

The mitochondrial network continually undergoes events of fission and fusion. Under physiologic conditions, the network is in equilibrium and is characterized by the presence of both elongated and punctate mitochondria. However, this balanced, homeostatic mitochondrial profile can change morphologic distribution in response to various stressors. Therefore, it is imperative to develop a method that robustly measures mitochondrial morphology with high accuracy. Here, we developed a semi-automated image analysis pipeline for the quantitation of mitochondrial morphology for both in vitro and in vivo applications. The image analysis pipeline was generated and validated utilizing images of primary cortical neurons from transgenic mice, allowing genetic ablation of key components of mitochondrial dynamics. This analysis pipeline was further extended to evaluate mitochondrial morphology in vivo through immunolabeling of brain sections as well as serial block-face scanning electron microscopy. These data demonstrate a highly specific and sensitive method that accurately classifies distinct physiological and pathological mitochondrial morphologies. Furthermore, this workflow employs the use of readily available, free open-source software designed for high throughput image processing, segmentation, and analysis that is customizable to various biological models.

The calpain family of proteases is causally linked to postischemic neurodegeneration. However, the precise mechanisms by which calpains contribute to postischemic neuronal death have not been fully elucidated. This review outlines the key features of the calpain system, and the evidence for its causal role in postischemic neuronal pathology. Furthermore, the consequences of specific calpain substrate cleavage at various subcellular locations are explored. Calpain substrates within synapses, plasma membrane, endoplasmic reticulum, lysosomes, mitochondria, and the nucleus, as well as the overall effect of postischemic calpain activity on calcium regulation and cell death signaling are considered. Finally, potential pathways for calpain-mediated neurodegeneration are outlined in an effort to guide future studies aimed at understanding the downstream pathology of postischemic calpain activity and identifying optimal therapeutic strategies.

Extracorporeal cardiopulmonary resuscitation (ECPR) is emerging as a feasible and effective rescue strategy for prolonged cardiac arrest (CA). However, prolonged total body ischemia and reperfusion can cause microvascular occlusion that prevents organ reperfusion and recovery of function. One hypothesized mechanism of microvascular “no-reflow” is leukocyte adhesion and formation of neutrophil extracellular traps. In this study we tested the hypothesis that a leukocyte filter (LF) or leukocyte modulation device (L-MOD) could reduce NETosis and improve recovery of heart and brain function in a swine model of prolonged cardiac arrest treated with ECPR. Thirty-six swine (45.5 ± 2.5 kg, evenly distributed sex) underwent 8 min of untreated ventricular fibrillation CA followed by 30 min of mechanical CPR with subsequent 8 h of ECPR. Two females were later excluded from analysis due to CPR complications. Swine were randomized to standard care (Control group), LF, or L-MOD at the onset of CPR. NET formation was quantified by serum dsDNA and citrullinated histone as well as immunofluorescence staining of the heart and brain for citrullinated histone in the microvasculature. Primary outcomes included recovery of cardiac function based on cardiac resuscitability score (CRS) and recovery of neurologic function based on the somatosensory evoked potential (SSEP) N20 cortical response. In this model of prolonged CA treated with ECPR we observed significant increases in serum biomarkers of NETosis and immunohistochemical evidence of microvascular NET formation in the heart and brain that were not reduced by LF or L-MOD therapy. Correspondingly, there were no significant differences in CRS and SSEP recovery between Control, LF, and L-MOD groups 8 h after ECPR onset (CRS = 3.1 ± 2.7, 3.7 ± 2.6, and 2.6 ± 2.6 respectively; p  = 0.606; and SSEP = 27.9 ± 13.0%, 36.7 ± 10.5%, and 31.2 ± 9.8% respectively, p  = 0.194). In this model of prolonged CA treated with ECPR, the use of LF or L-MOD therapy during ECPR did not reduce microvascular NETosis or improve recovery of myocardial or brain function. The causal relationship between microvascular NETosis, no-reflow, and recovery of organ function after prolonged cardiac arrest treated with ECPR requires further investigation.

Intranasal insulin is a putative neuroprotective therapy after cardiac arrest, but safety in humans at doses extrapolated from animal models is unknown. This phase I, open‐label adaptive dose‐escalation study explores the maximum tolerated dose of intranasal insulin in healthy human participants. Placebo or insulin at doses from 0 to 1000 units was given to healthy participants intranasally on repeated weekly visits. Serum glucose, insulin, and C‐peptide levels were measured serially at 0, 15, 30, 60, 120, 180, and 240 min after administration. Twenty‐four participants (12 female, median age 53, IQR 35–61) were enrolled. There was minimal change in average serum glucose after administration of intranasal insulin. Average serum insulin increased slightly in a dose‐dependent manner, reaching maximum concentrations at 15 min. C‐peptide decreased over time from administration in all groups. One participant had severe hypoglycemia (24 mg/dL at 45 min) and a different participant had mild hypoglycemia (51 mg/dL at 30 min), both after receiving 600 U intranasal insulin. Hypoglycemic episodes were associated with increases in serum insulin. Both participants continued in the study without hypoglycemia after additional doses. High‐dose intranasal insulin up to 1000 U was generally well tolerated, with minimal measurable systemic absorption and without significant aggregate changes in mean glucose. Idiosyncratic episodic systemic absorption and hypoglycemia require further study and additional caution in potential clinical application. Further study of its target engagement and efficacy as a neuroprotective therapy after cardiac arrest at these doses is warranted.

As more people are surviving cardiac arrest, focus needs to shift towards improving neurological outcomes and quality of life in survivors. Brain injury after resuscitation, a common sequela following cardiac arrest, ranges in severity from mild impairment to devastating brain injury and brainstem death. Effective strategies to minimise brain injury after resuscitation include early intervention with cardiopulmonary resuscitation and defibrillation, restoration of normal physiology, and targeted temperature management. It is important to identify people who might have a poor outcome, to enable informed choices about continuation or withdrawal of life-sustaining treatments. Multimodal prediction guidelines seek to avoid premature withdrawal in those who might survive with a good neurological outcome, or prolonging treatment that might result in survival with severe disability. Approximately one in three admitted to intensive care will survive, many of whom will need intensive, tailored rehabilitation after discharge to have the best outcomes.

Purpose We sought to generate national estimates for post-cardiac arrest mortality, to assess trends, and to identify hospital factors associated with survival. Methods We used a national sample of US hospitals to identify patients resuscitated after cardiac arrest from 2000 to 2004 to describe the association between hospital factors (teaching status, location, size) and mortality, length of stay, and hospital charges. Analyses were performed using logistic regression. Results A total of 109,739 patients were identified. In-hospital mortality was 70.6%. A 2% decrease in unadjusted mortality from 71.6% in 2000 to 69.6% in 2004 (OR 0.96, P  < 0.001) was observed. Mortality was lower at teaching hospitals (OR 0.58, P  = 0.001), urban hospitals (OR 0.63, P  = 0.004), and large hospitals (OR 0.55, P  < 0.001). Conclusion Mortality after in-hospital cardiac arrest decreased over 5 years. Mortality was lower at urban, teaching, and large hospitals. There are implications for dissemination of best practices or regionalization of post-cardiac arrest care.

Development of an accurate system to predict the daily incidence of out-of-hospital cardiac arrest (OHCA) might provide a significant public health benefit. We developed a machine learning model to predict daily OHCA incidence at the regional level using high-resolution meteorological data, chronological data, and sociodemographic data. Among OHCAs of non-traumatic cause, 196,735 in the training dataset (2013-2017), 119,455 in the testing dataset (2018-2019) from internal areas, and 37,160 in the testing dataset from external areas were included in the analysis. Application of invariant causal prediction (ICP) successfully reduced the number of variables to 17 while maintaining high predictive performance for nationwide incidence per 100,000 per day in both the training and testing datasets, whether from internal or external regions, comparable to the non-ICP model using the initial 34 variables. Furthermore, the prediction model retained a satisfactory performance up to seven days in advance.

ImportanceValue in health care is quality per unit cost (V = Q/C), and an emergency department–based intensive care unit (ED-ICU) model has been associated with improved quality. To assess the value of this care delivery model, it is essential to determine the incremental direct cost of care.ObjectiveTo determine the association of an ED-ICU with inflation-adjusted change in mean direct cost of care, net revenue, and direct margin per ED patient encounter.Design, Setting, and ParticipantsThis retrospective economic analysis evaluated the cost of care delivery to patients in the ED before and after deployment of the Joyce and Don Massey Family Foundation Emergency Critical Care Center, an ED-ICU, on February 16, 2015, at a large academic medical center in the US with approximately 75 000 adult ED visits per year. The pre–ED-ICU cohort was defined as all documented ED visits by patients 18 years or older with a complete financial record from September 8, 2012, through June 30, 2014 (660 days); the post–ED-ICU cohort, all visits from July 1, 2015, through April 21, 2017 (660 days). Fiscal year 2015 was excluded from analysis to phase in the new care model. Statistical analysis was performed March 1 through December 30, 2021.ExposuresImplementation of an ED-ICU.Main Outcomes and MeasuresInflation-adjusted direct cost of care, net revenue, and direct margin per patient encounter in the ED.ResultsA total of 234 884 ED visits during the study period were analyzed, with 115 052 patients (54.7% women) in the pre–ED-ICU cohort and 119 832 patients (54.5% women) in the post–ED-ICU cohort. The post–ED-ICU cohort was older (mean [SD] age, 49.1 [19.9] vs 47.8 [19.6] years;P < .001), required more intensive respiratory support (2.2% vs 1.1%;P < .001) and more vasopressor use (0.5% vs 0.2%;P < .001), and had a higher overall case mix index (mean [SD], 1.7 [2.0] vs 1.5 [1.7];P < .001). Implementation of the ED-ICU was associated with similar inflation-adjusted total direct cost per ED encounter (pre–ED-ICU, mean [SD],$4875 [$ 15 175]; post–ED-ICU,$4877 [$ 17 400];P = .98). Inflation-adjusted net revenue per encounter increased by 7.0% (95% CI, 3.4%-10.6%;P < .001), and inflation-adjusted direct margin per encounter increased by 46.6% (95% CI, 32.1%-61.2%;P < .001).Conclusions and RelevanceImplementation of an ED-ICU was associated with no significant change in inflation-adjusted total direct cost per ED encounter. Holding delivery costs constant while improving quality demonstrates improved value via the ED-ICU model of care.

IntroductionOut-of-hospital cardiac arrest (OHCA) is a common, life-threatening event encountered routinely by first responders, including police, fire and emergency medical services (EMS). Current literature suggests that there is significant regional variation in outcomes, some of which may be related to modifiable factors. Yet, there is a persistent knowledge gap regarding strategies to guide quality improvement efforts in OHCA care and, by extension, survival. The Enhancing Prehospital Outcomes for Cardiac Arrest (EPOC) study aims to fill these gaps and to improve outcomes.Methods and analysisThis mixed-methods study includes three aims. In aim I, we will define variation in OHCA survival to the emergency department (ED) among EMS agencies that participate in the Michigan Cardiac Arrest Registry to Enhance Survival (CARES) in order to sample EMS agencies with high-survival and low-survival outcomes. In aim II, we will conduct site visits to emergency medical systems—including 911/dispatch, police, non-transport fire, and EMS agencies—in approximately eight high-survival and low-survival communities identified in aim I. At each site, key informant interviews and a multidisciplinary focus group will identify themes associated with high OHCA survival. Transcripts will be coded using a structured codebook and analysed through thematic analysis. Results from aims I and II will inform the development of a survey instrument in aim III that will be administered to all EMS agencies in Michigan. This survey will test the generalisability of factors associated with increased OHCA survival in the qualitative work to ultimately build an EPOC Toolkit which will be distributed to a broad range of stakeholders as a practical ‘how-to’ guide to improve outcomes.Ethics and disseminationThe EPOC study was deemed exempt by the University of Michigan Institutional Review Board. Findings will be compiled in an ‘EPOC Toolkit’ and disseminated in the USA through partnerships including, but not limited to, policymakers, EMS leadership and health departments.