Quality improvement developments following pediatric resuscitation and veno-arterial extracorporeal membrane oxygenation support due to a massive intentional antidepressant overdose.

Extracorporeal Membrane Oxygenation Cardiopulmonary Resuscitation (ECPR) is the act of placing a patient on bypass at the bedside while simultaneously carrying out life-sustaining interventions such as chest compressions or epinephrine administration. This involves a team of physicians, nurses, respiratory therapists, pharmacists, extracorporeal membrane oxygenation (ECMO) trained staff, and other health professionals who must focus on cardiopulmonary resuscitation (CPR), cannulation, and initiating ECMO flow at the same time. ECPR may be considered when traditional CPR does not achieve return of spontaneous circulation (ROSC) in a patient. Limitations when thinking about using ECPR for a patient include location, timing from arrest to CPR initiation, as well as CPR initiation to successfully on bypass, trained staff available to begin the cannulation process, and pauses in compressions during surgery. We analyzed a pediatric patient who required ECPR after an intentional drug overdose. Gaps identified in this case prompted us to assess our ECPR protocol. Through the development and use of multidisciplinary ECPR simulations, our team discovered areas of quality improvement and put those findings into practice.

Melanomas with concurrent BRAF non-p.V600 and NF1 loss-of-function mutations are targetable by BRAF/MEK inhibitor combination therapy.

Although combination BRAF/MEK inhibition has produced significant survival benefits for BRAF p.V600 mutant melanomas, targeted therapies approved for BRAF non-p.V600 mutant melanomas remain limited. Through the analysis of 772 cutaneous melanoma exomes, we reveal that BRAF non-p.V600 mutations co-occurs more frequently with NF1 loss, but not with oncogenic NRAS mutations, than expected by chance. We present cell signaling data, which demonstrate that BRAF non-p.V600 mutants can signal as monomers and dimers within an NF1 loss context. Concordantly, BRAF inhibitors that inhibit both monomeric and dimeric BRAF synergize with MEK inhibition to significantly reduce cell viability in vitro and tumor growth in vivo in BRAF non-p.V600 mutant melanomas with co-occurring NF1 loss-of-function mutations. Our data suggest that patients harboring BRAF non-p.V600 mutant melanomas may benefit from current FDA-approved BRAF/MEK inhibitor combination therapy currently reserved for BRAF p.V600 mutant patients.

Spatially mapping the immune landscape of melanoma using imaging mass cytometry.

Melanoma is an immunogenic cancer with a high response rate to immune checkpoint inhibitors (ICIs). It harbors a high mutation burden compared with other cancers and, as a result, has abundant tumor-infiltrating lymphocytes (TILs) within its microenvironment. However, understanding the complex interplay between the stroma, tumor cells, and distinct TIL subsets remains a substantial challenge in immune oncology. To properly study this interplay, quantifying spatial relationships of multiple cell types within the tumor microenvironment is crucial. To address this, we used cytometry time-of-flight (CyTOF) imaging mass cytometry (IMC) to simultaneously quantify the expression of 35 protein markers, characterizing the microenvironment of 5 benign nevi and 67 melanomas. We profiled more than 220,000 individual cells to identify melanoma, lymphocyte subsets, macrophage/monocyte, and stromal cell populations, allowing for in-depth spatial quantification of the melanoma microenvironment. We found that within pretreatment melanomas, the abundance of proliferating antigen-experienced cytotoxic T cells (CD8+CD45RO+Ki67+) and the proximity of antigen-experienced cytotoxic T cells to melanoma cells were associated with positive response to ICIs. Our study highlights the potential of multiplexed single-cell technology to quantify spatial cell-cell interactions within the tumor microenvironment to understand immune therapy responses.

Endothelial biology in humans: from acetylcholine testing in coronary arteries to personalized management of coronary heart disease.

The role of acetylcholine in determining the tone of blood vessels.

Guy Fontaine and arrhythmogenic right ventricular dysplasia.

Cardiomyopathies are the Focus of this EHJ issue. Guy Fontaine, an electrical engineer, then a physician, played a pivotal role in the discovery of ARVD.

Polymorphic variation in cytochrome oxidase subunit genes.

Cytochrome oxidase (COX) activity varies between individuals and low activities associate with Alzheimer's disease. Whether genetic heterogeneity influences function of this multimeric enzyme is unknown. To explore this we sequenced three mitochondrial DNA (mtDNA) and ten nuclear COX subunit genes from at least 50 individuals. 20% had non-synonymous mtDNA COX gene polymorphisms, 12% had a COX4I1 non-synonymous G to A transition, and other genes rarely contained non-synonymous polymorphisms. Frequent untranslated region (UTR) polymorphisms were seen in COX6A1, COX6B1, COX6C, and COX7A1; heterogeneity in a COX7A1 5' UTR Sp1 site was extensive. Synonymous polymorphisms were common and less frequent in the more conserved COX1 than the less conserved COX3, suggesting at least in mtDNA synonymous polymorphisms experience selection pressure and are not functionally silent. Compound gene variations occurred within individuals. To test whether variations could have functional consequences, we studied the COX4I1 G to A transition and an AGCCCC deletion in the COX7A1 5' UTR Sp1 site. Cells expressing the COX4I1 polymorphism had reduced COX Vmax activity. In reporter construct-transduced cells where green fluorescent protein expression depended on the COX7A1 Sp1 site, AGCCCC deletion reduced fluorescence. Our findings indicate COX subunit gene heterogeneity is pervasive and may mediate COX functional variation.

Effects of memantine on mitochondrial function.

Because NMDA complex and mitochondrial function are related, we hypothesized memantine would influence mitochondrial function. We addressed this in vitro by studying the effects of chronic and acute memantine exposures on mitochondrial function. For acute exposure experiments, mitochondria were isolated from NT2 cells and assayed for electron transport chain (ETC) enzyme function and peroxide production in buffers containing up to 60uM memantine. For chronic exposure experiments, NT2 cells were maintained for at least two weeks in medium containing up to 60uM memantine, following which we assayed cells or their mitochondria for ETC enzyme activities, cytochrome oxidase protein levels, oxidative stress, calcium levels, and mitochondrial DNA levels. The ability of the NMDA receptor antagonist aminophosphonovaleric acid (APV) to modify memantine's mitochondrial effects was evaluated. Acute and chronic memantine similarly affected complex I (increased at high concentrations) and IV (decreased at high concentrations) V(max) activities. APV did not alter the effects of chronic memantine exposure on citrate synthase and complex IV. We detected a lower mitochondrial peroxide production rate with acute exposure, and an increased mitochondrial peroxide production rate with chronic exposure. Micromolar memantine concentrations affect mitochondria, some of these effects are directly mediated, and acute and chronic effects may differ.