Publications

INTRODUCTION: Ubiquinol (reduced coenzyme Q10) is essential for adequate aerobic metabolism. The objective of this trial was to determine whether ubiquinol administration in patients resuscitated from cardiac arrest could increase physiological coenzyme Q10 levels, improve oxygen consumption, and reduce neurological biomarkers of injury.

MATERIALS AND METHODS: This was a randomized, double-blind, placebo-controlled trial in patients successfully resuscitated from cardiac arrest. Patients were randomized to receive enteral ubiquinol (300 mg) or placebo every 12 h for up to 7 days. The primary endpoint was total coenzyme Q10 plasma levels at 24 h after enrollment. Secondary endpoints included neuron specific enolase, S100B, lactate, cellular and global oxygen consumption, neurological status, and in-hospital mortality.

RESULTS: Forty-three patients were included in the modified intention-to-treat analysis. Median coenzyme Q10 levels were significantly higher in the ubiquinol group as compared to the placebo group at 24 h (441 [IQR, 215-510] ηg/mL vs. 113 [IQR, 94-208] ηg/mL, P < 0.001). Similar results were observed at 48 and 72 h. There were no differences between the two groups in any of the secondary endpoints. Median neuron specific enolase levels were not different between the two groups at 24 h (16.8 [IQR, 9.5-19.8] ηg/mL vs. 8.2 [IQR, 4.3-19.1] ηg/mL, P = 0.61).

CONCLUSIONS: Administration of enteral ubiquinol increased plasma coenzyme Q10 levels in post-cardiac arrest patients as compared to placebo. There were no differences in neurological biomarkers and oxygen consumption between the two groups.

Background: BRAFV600E acts as an ATP-dependent cytosolic kinase. BRAFV600E inhibitors are widely available, but resistance to them is widely reported in the clinic. Lipid metabolism (fatty acids) is fundamental for energy and to control cell stress. Whether and how BRAFV600E impacts lipid metabolism regulation in papillary thyroid carcinoma (PTC) is still unknown. Acetyl-CoA carboxylase (ACC) is a rate-limiting enzyme for de novo lipid synthesis and inhibition of fatty acid oxidation (FAO). ACC1 and ACC2 genes encode distinct isoforms of ACC. The aim of our study was to determine the relationship between BRAFV600E and ACC in PTC. Methods: We performed RNA-seq and DNA copy number analyses in PTC and normal thyroid (NT) in The Cancer Genome Atlas samples. Validations were performed by using assays on PTC-derived cell lines of differing BRAF status and a xenograft mouse model derived from a heterozygous BRAFWT/V600E PTC-derived cell line with knockdown (sh) of ACC1 or ACC2. Results:ACC2 mRNA expression was significantly downregulated in BRAFV600E-PTC vs. BRAFWT-PTC or NT clinical samples. ACC2 protein levels were downregulated in BRAFV600E-PTC cell lines vs. the BRAFWT/WT PTC cell line. Vemurafenib increased ACC2 (and to a lesser extent ACC1) mRNA levels in PTC-derived cell lines in a BRAFV600E allelic dose-dependent manner. BRAFV600E inhibition increased de novo lipid synthesis rates, and decreased FAO due to oxygen consumption rate (OCR), and extracellular acidification rate (ECAR), after addition of palmitate. Only shACC2 significantly increased OCR rates due to FAO, while it decreased ECAR in BRAFV600E PTC-derived cells vs. controls. BRAFV600E inhibition synergized with shACC2 to increase intracellular reactive oxygen species production, leading to increased cell proliferation and, ultimately, vemurafenib resistance. Mice implanted with a BRAFWT/V600E PTC-derived cell line with shACC2 showed significantly increased tumor growth after vemurafenib treatment, while vehicle-treated controls, or shGFP control cells treated with vemurafenib showed stable tumor growth. Conclusions: These findings suggest a potential link between BRAFV600E and lipid metabolism regulation in PTC. BRAFV600E downregulates ACC2 levels, which deregulates de novo lipid synthesis, FAO due to OCR, and ECAR rates. ShACC2 may contribute to vemurafenib resistance and increased tumor growth. ACC2 rescue may represent a novel molecular strategy for overcoming resistance to BRAFV600E inhibitors in refractory PTC.

UNLABELLED: Lower oxygen consumption is associated with worse survival in septic shock and in other forms of critical illness. No treatment that increases oxygen extraction, a key determinant of oxygen consumption, has been found. Thiamine is required for aerobic metabolism, and deficiency is common in the critically ill.

OBJECTIVES: We evaluated the effect of thiamine on oxygen consumption in patients requiring mechanical ventilation for an acute illness.

DESIGN: Phase II, randomized, double-blind, and placebo-controlled trial.

SETTING AND PARTICIPANTS: ICUs in a tertiary care hospital in the United States. Patients admitted to the ICU and requiring mechanical ventilation were screened for enrollment.

INTERVENTIONS: After enrollment, baseline measurement of oxygen consumption and baseline laboratories including lactate, central venous oxygen saturation, and pyruvate dehydrogenase, a single dose of 200 mg IV thiamine or placebo was administered. Oxygen consumption was then monitored for 6 additional hours and repeat laboratories were drawn at the end of the protocol.

MAIN OUTCOMES AND MEASURES: The primary outcome was the change in oxygen consumption. Analysis was done using linear regression with a first-order autoregressive variance-covariance structure to account for repeated measures within subjects. Secondary outcomes included change in lactate, central venous oxygen saturation, and pyruvate dehydrogenase quantity and activity.

RESULTS: Sixty-seven patients were enrolled. After excluding 11 patients due to inadequate quantity or quality of oxygen consumption data, 56 patients were included. There was no difference in change in oxygen consumption in the 6 hours after study drug. Results for secondary outcomes were similarly negative. In the prespecified subgroup of 18 thiamine deficient patients, there was a difference in the two oxygen consumption curves (p = 0.006), although no difference in median oxygen consumption or area under the curve.

CONCLUSIONS AND RELEVANCE: A single dose of IV thiamine did not alter oxygen consumption in patients requiring mechanical ventilation for acute illness.

UNLABELLED: mRNA vaccines can be developed and produced quickly, making them attractive for immediate outbreak responses. Furthermore, clinical trials have demonstrated rapid protection following mRNA vaccination. We sought to investigate how quickly mRNA vaccines elicit antibody responses compared to other vaccine modalities. We first examined immune kinetics of mRNA and DNA vaccines expressing SARS-CoV-2 spike in mice. We observed rapid induction of antigen-specific binding and neutralizing antibodies by day 5 following mRNA, but not DNA, immunization. The mRNA vaccine also induced increased levels of IL-5, IL-6 and MCP-1. We then evaluated immune kinetics of an HIV-1 mRNA vaccine in comparison to DNA, protein, and rhesus adenovirus 52 (RhAd52) vaccines with the same HIV-1 envelope antigen in mice. Induction of envelope-specific antibodies was observed by day 5 following mRNA vaccination, whereas antibodies were detected by day 7-14 following DNA, protein, and RhAd52 vaccination. Eliciting rapid humoral immunity may be an advantageous property of mRNA vaccines for controlling infectious disease outbreaks.

IMPORTANCE: mRNA vaccines can be developed and produced in record time. Here we demonstrate induction of rapid antibody responses by mRNA vaccines encoding two different viral antigens by day 5 following immunization in mice. The rapid immune kinetics of mRNA vaccines can be an advantageous property that makes them well suited for rapid control of infectious disease outbreaks.

Ubiquinol is a fundamental component of cellular metabolism. Low ubiquinol levels have been associated with mortality. This was a substudy of a randomized trial in patients undergoing coronary artery bypass grafting. We drew blood before and after surgery. Ubiquinol or placebo was added to peripheral blood mononuclear cells for oxygen consumption (OCR) measurements. In vivo ubiquinol levels were lower postsurgery compared to presurgery (0.16 μmol/L [quartiles: 0.02-0.39], P = .01), although the difference disappeared when adjusting for hemoglobin levels (P = .30). There was no difference in presurgical basal (1.0 mL/min/mg [95% confidence interval [CI]: -0.9 to 2.2], P = .08) and maximal (0.5 mL/min/mg [95% CI: -4.3 to 7.3], P = .56) OCR in cells receiving ubiquinol or placebo. There was a difference in postsurgical basal (1.1 mL/min/mg [95% CI: 0.9-1.6], P < .001) and maximal (4.2 mL/min/mg [95% CI: 0.3-7.0], P = .01) OCR between the groups. We found no association between ubiquinol and OCR levels (all P > .05).

INTRODUCTION: Vitamin C deficiency has been described in patients with sepsis. The post-cardiac arrest syndrome shares similarities to sepsis, however vitamin C levels in post-arrest patients have been incompletely characterized. We assessed vitamin C levels in a post-arrest population.

METHODS: This was a retrospective observational study at a tertiary care center. A convenience sample of post-arrest, sepsis, and healthy control patients was selected from prior studies. Vitamin C levels were measured from samples obtained within 6-h of emergency department admission. A subset of cardiac arrest patients had vitamin C levels additionally measured 24-h later.

RESULTS: A total of 84 patients (34 healthy controls, 25 post-arrest, and 25 septic patients) were included. The median baseline vitamin C level in cardiac arrest patients was 0.33 mg/dL (0.05-0.83), as compared to 0.91 mg/dL (0.69-1.48) in the healthy control group (p < 0.01) and 0.28 mg/dL (0.11-0.59) in the septic group (p = 0.36). Vitamin C levels for cardiac arrest patients fell between the two time points, but the change was not statistically significant (median decrease 0.26 mg/dL, p = 0.08).

CONCLUSIONS: Serum vitamin C levels were lower in post-arrest patients compared to controls and were similar to patients with sepsis. Future studies of vitamin C levels and supplementation following cardiac arrest may be warranted.

Background Neuromuscular blockade (NMB) agents are often administered to control shivering during targeted temperature management following cardiac arrest. In this study, we hypothesized that early, continuous NMB would result in a greater reduction in serum lactate levels among comatose patients after cardiac arrest. Methods and Results Randomized trial of continuous NMB for 24 hours versus usual care following cardiac arrest conducted at 5 urban centers in the United States. Adult patients who achieved return of spontaneous circulation, remained unresponsive, and underwent targeted temperature management after cardiac arrest were included. The primary outcome was change in lactate over 24 hours. A total of 83 patients were randomized, and 80 were analyzed (37 and 43 in the NMB and usual care arms, respectively). There was no significant interaction between time and treatment group with respect to change in lactate over 24 hours (median lactate change from 4.2 to 2.0 mmol/L [-2.2 mmol/L] in the NMB arm versus 4.0 to 1.7 mmol/L [-2.3 mmol/L] in the usual care arm; geometric mean difference, 1.3 [95% CI, 1.0-1.8]; P=0.07 for the interaction term). There was no difference in hospital survival (38% [NMB] versus 33% [usual care]; P=0.63) or survival with good functional outcome (30% [NMB] versus 21% [usual care]; P=0.35). There were no adverse events in either arm attributed to study interventions. Conclusions Continuous NMB compared with usual care did not reduce lactate over the first 24 hours after enrollment compared with usual care. There was no difference in overall hospital survival, hospital survival with good neurologic outcome, or adverse events. Registration URL: https://www.clinicaltrials.gov; Unique identifier: NCT02260258.

BACKGROUND: Outcome prediction after out-of-hospital cardiac arrest (OHCA) is difficult. We hypothesized that lactate and need for vasopressors would predict outcome, and that addition of a mitochondrial biomarker would enhance performance of the tool.

METHODS: Prospective observational study of OHCA patients presenting to an academic medical center September 2008 to April 2016. We conducted univariate and multivariate logistic regressions.

RESULTS: Patients were divided based on 2 variables: vasopressor status and initial lactate (<5 mmol/L, 5-10, ≥10). Three hundred fifty-two patients were evaluated; 249 had a lactate within 3 hours and were included. Patients on vasopressors had higher mortality (74% vs 40%; P < .001). A stepwise increase in mortality is associated with increasing lactate (45% lactate <5, 66% 5-10, and 83% ≥10; P < 001). Multivariable models with lactate group and vasopressors as predictors demonstrated excellent discrimination (area under the receiver operating curve [AUC]: 0.73 [95% confidence interval, CI: 0.66-0.79]; adjusted for additional covariates: AUC: 0.81 [95% CI: 0.75-0.86]). Thirty-six patients had cytochrome c levels available; among these 36, when comparing models with and without cytochrome c, there was no difference (AUC: 0.88 [95% CI: 0.76-1.00] vs AUC: 0.85 [95% CI: 0.73-0.98], respectively; P = .30).

CONCLUSION: In this prospective validation, the combination of lactate and vasopressors in the immediate postarrest period is predictive of mortality. Cytochrome c offered minimal additional predictive power.

OBJECTIVES: The goal of this investigation was to determine if acute influenza infection is associated with depletion of CoQ10 compared to healthy controls and to determine any associations between CoQ10 levels and illness severity and inflammatory biomarkers.

PATIENTS AND METHODS: We analyzed serum CoQ10 concentrations of patients with acute influenza enrolled in a randomized clinical trial prior to study drug administration. Patients were enrolled at a single urban tertiary care center over 3 influenza seasons (December 27, 2013 to March 31, 2016). Wilcoxon rank sum test was used to compare CoQ10 levels between influenza patients and healthy controls. Correlations with inflammatory biomarkers and severity of illness were assessed using Spearman correlation coefficient.

RESULTS: We analyzed CoQ10 levels from 50 patients with influenza and 29 controls. Overall, patients with acute influenza had lower levels of CoQ10 (.53 μg/mL, IQR .37-.75 vs .72, IQR .58-.90, P = .004). Significantly more patients in the influenza group had low CoQ10 levels (<.5 μg/mL) compared to controls (48% vs 7%, P < .001). Among influenza patients, there were significant but weak correlations between CoQ10 levels and IL-2 (r = -.30, P = .04), TNF-alpha (r = -.35, P = .01) and VEGF (r = .38, P = .007), but no correlation with IL-6, IL-10, VCAM or influenza severity of illness score (all P > .05).

CONCLUSIONS: We found that CoQ10 levels were significantly lower in patients with acute influenza infection and that these levels had significant although weak correlations with several inflammatory biomarkers.

BACKGROUND: The biological effects of nitric oxide are mediated via protein S-nitrosylation. Levels of S-nitrosylated protein are controlled in part by the denitrosylase, S-nitrosoglutathione reductase (GSNOR). The objective of this study was to examine whether GSNOR inhibition improves outcomes after cardiac arrest and cardiopulmonary resuscitation (CA/CPR).

METHODS: Adult wild-type C57BL/6 and GSNOR-deleted (GSNOR-/-) mice were subjected to potassium chloride-induced CA and subsequently resuscitated. Fifteen minutes after a return of spontaneous circulation, wild-type mice were randomized to receive the GSNOR inhibitor, SPL-334.1, or normal saline as placebo. Mortality, neurological outcome, GSNOR activity, and levels of S-nitrosylated proteins were evaluated. Plasma GSNOR activity was measured in plasma samples obtained from post-CA patients, preoperative cardiac surgery patients, and healthy volunteers.

RESULTS: GSNOR activity was increased in plasma and multiple organs of mice, including brain in particular. Levels of protein S-nitrosylation were decreased in the brain 6 hours after CA/CPR. Administration of SPL-334.1 attenuated the increase in GSNOR activity in brain, heart, liver, spleen, and plasma, and restored S-nitrosylated protein levels in the brain. Inhibition of GSNOR attenuated ischemic brain injury and improved survival in wild-type mice after CA/CPR (81.8% in SPL-334.1 versus 36.4% in placebo; log rank P=0.031). Similarly, GSNOR deletion prevented the reduction in the number of S-nitrosylated proteins in the brain, mitigated brain injury, and improved neurological recovery and survival after CA/CPR. Both GSNOR inhibition and deletion attenuated CA/CPR-induced disruption of blood brain barrier. Post-CA patients had higher plasma GSNOR activity than did preoperative cardiac surgery patients or healthy volunteers ( P<0.0001). Plasma GSNOR activity was positively correlated with initial lactate levels in postarrest patients (Spearman correlation coefficient=0.48; P=0.045).

CONCLUSIONS: CA and CPR activated GSNOR and reduced the number of S-nitrosylated proteins in the brain. Pharmacological inhibition or genetic deletion of GSNOR prevented ischemic brain injury and improved survival rates by restoring S-nitrosylated protein levels in the brain after CA/CPR in mice. Our observations suggest that GSNOR is a novel biomarker of postarrest brain injury as well as a molecular target to improve outcomes after CA.