Identifying Circulating and Lung Tissue Cytokines Associated with Thoracic Irradiation and AEOL 10150 Treatment in a Nonhuman Primate Model.

Inflammatory cytokines have been suggested to play important roles in radiation-induced lung injury (RILI). Identifying significantly changed circulating and tissue cytokines after thoracic irradiation will aid in deciphering the mechanism of RILI and identifying potential biomarkers to predict clinical outcome. Herein, the levels of 24 cytokines were measured in serial plasma samples and lung tissue samples collected from a pilot study where nonhuman primates (NHPs) received 11.5 Gy whole thoracic lung irradiation (WTLI) and were then treated with or without a medical countermeasure, AEOL 10150 [a superoxide dismutase (SOD) mimetic]. Seven plasma cytokines (i.e., IP-10, MCP-1, IL-12, IL-15, IL-16, IL-7 and IL-6) were found to be significantly changed at different time points due to WTLI. Plasma IP-10 and MDC were significantly changed between the vehicle group and the drug group. The levels of IP-10, MCP-1, MIP-1α, TARC, IL-17, TNF-ß and IL-6 were significantly elevated in the lung tissue lysates of NHPs that received WTLI versus radiation-naïve NHPs. The terminal plasma concentrations of IP-10, MDC, TARC, IL-12, IL-15 and IL-6 were significantly correlated with their levels in the lung tissue. The levels of four cytokines (MCP-4, IL-17, TNF-ß and IL-2) at early time points (≤8 weeks postirradiation) were significantly correlated with their terminal plasma levels, respectively. Statistical analysis indicated that circulating cytokines could be discriminatory predictors of AEOL 10150 treatment. Taken together, our data suggested that the cytokine profiles were significantly changed after WTLI as well as mitigator treatment, and that the plasma cytokine profiles could potentially be used to distinguish vehicle or mitigator treatment after WTLI in a NHP model.

Pyruvate-enriched resuscitation for shock.

This commentary addresses the recent retraction of an article which reported favorable outcomes in septic patients treated with intravenous pyruvate. The retracted report was cited in the authors' recent minireview on the cellular mechanisms and clinical application of pyruvate to improve cardiac performance. Because the retracted article reports pyruvate-enhanced resuscitation of critically ill patients, the authors wish to inform the readership, especially critical care providers, that this particular clinical application of pyruvate is not now supported by robust evidence. After discussing the retraction's implications for the clinical application of pyruvate-enriched resuscitation for sepsis, this commentary summarizes the extensive preclinical evidence of the efficacy and mechanisms of pyruvate resuscitation in animal models of hemorrhagic and septic shock, which argues for renewed clinical investigation of pyruvate-enriched resuscitation. Impact statement This commentary addresses the recent retraction of a clinical report of significant benefits of intravenous pyruvate resuscitation in septic patients, including sharply lowered mortality and decreased circulating pro-inflammatory cytokines, which was cited in the authors' minireview in Experimental Biology and Medicine. The potential implications of the retraction, and the extensive preclinical evidence supporting the use of pyruvate-enriched resuscitation for shock states, are summarized and discussed.

Effect of Norepinephrine on Intracellular Ca2+ Levels in Malignant Hyperthermia-Susceptible B Cells: Pilot Study in the Search for a New Diagnostic Test for Malignant Hyperthermia.

Malignant hyperthermia (MH) crises may induce morbidity or death in MH-susceptible (MHS) individuals. The only sensitive method of determining susceptibility is the caffeine-halothane contracture test, requiring muscle biopsy. Early research on MH demonstrated an abnormal response to catecholamines in MHS individuals. The purpose of this study was to determine whether MHS B lymphocytes would demonstrate an increased sensitivity to norepinephrine as indicated by an adrenergic augmentation of intracellular calcium ion (Ca2+) accumulation, to possibly develop a less invasive laboratory assay for determining MH susceptibility. The fluorescent Ca2+ indicator dye fura-2 acetoxymethyl was used to identify Ca2+ flux within Epstein-Barr virus- immortalized MH-negative (MHN) and MHS B cells exposed to the RyR1 agonist 4-chloro-m-cresol (4-CmC) before and after administration of 1 µM of norepinephrine. In the presence of 4-CmC and norepinephrine, the area under the curve dose responses were significantly elevated in MHS B cells compared with MHN B cells (F[1,10] = 27.37; P < .01). Epstein-Barr virus-immortalized B cells from MHS humans displayed an increased sensitivity to norepinephrine compared with those from MHN individuals. These data suggest that an abnormal response to exogenous norepinephrine could potentially be used to develop a diagnostic laboratory assay to determine MH susceptibility.

Pyruvate enhancement of cardiac performance: Cellular mechanisms and clinical application.

Cardiac contractile function is adenosine-5'-triphosphate (ATP)-intensive, and the myocardium's high demand for oxygen and energy substrates leaves it acutely vulnerable to interruptions in its blood supply. The myriad cardioprotective properties of the natural intermediary metabolite pyruvate make it a potentially powerful intervention against the complex injury cascade ignited by myocardial ischemia-reperfusion. A readily oxidized metabolic substrate, pyruvate augments myocardial free energy of ATP hydrolysis to a greater extent than the physiological fuels glucose, lactate and fatty acids, particularly when it is provided at supra-physiological plasma concentrations. Pyruvate also exerts antioxidant effects by detoxifying reactive oxygen and nitrogen intermediates, and by increasing nicotinamide adenine dinucleotide phosphate reduced form (NADPH) production to maintain glutathione redox state. These enhancements of free energy and antioxidant defenses combine to augment sarcoplasmic reticular Ca2+ release and re-uptake central to cardiac mechanical performance and to restore ß-adrenergic signaling of ischemically stunned myocardium. By minimizing Ca2+ mismanagement and oxidative stress, pyruvate suppresses inflammation in post-ischemic myocardium. Thus, pyruvate administration stabilized cardiac performance, augmented free energy of ATP hydrolysis and glutathione redox systems, and/or quelled inflammation in a porcine model of cardiopulmonary bypass, a canine model of cardiac arrest-resuscitation, and a caprine model of hypovolemia and hindlimb ischemia-reperfusion. Pyruvate's myriad benefits in preclinical models provide the mechanistic framework for its clinical application as metabolic support for myocardium at risk. Phase one trials have demonstrated pyruvate's safety and efficacy for intravenous resuscitation for septic shock, intracoronary infusion for heart failure and as a component of cardioplegia for cardiopulmonary bypass. The favorable outcomes of these trials, which argue for expanded, phase three investigations of pyruvate therapy, mirror findings in isolated, perfused hearts, underscoring the pivotal role of preclinical research in identifying clinical interventions for cardiovascular diseases. Impact statement This article reviews pyruvate's cardioprotective properties as an energy-yielding metabolic fuel, antioxidant and anti-inflammatory agent in mammalian myocardium. Preclinical research has shown these properties make pyruvate a powerful intervention to curb the complex injury cascade ignited by ischemia and reperfusion. In ischemically stunned isolated hearts and in large mammal models of cardiopulmonary bypass, cardiac arrest-resuscitation and hypovolemia, intracoronary pyruvate supports recovery of myocardial contractile function, intracellular Ca2+ homeostasis and free energy of ATP hydrolysis, and its antioxidant actions restore ß-adrenergic signaling and suppress inflammation. The first clinical trials of pyruvate for cardiopulmonary bypass, fluid resuscitation and intracoronary intervention for congestive heart failure have been reported. Receiver operating characteristic analyses show remarkable concordance between pyruvate's beneficial functional and metabolic effects in isolated, perfused hearts and in patients recovering from cardiopulmonary bypass in which they received pyruvate- vs. L-lactate-fortified cardioplegia. This research exemplifies the translation of mechanism-oriented preclinical studies to clinical application and outcomes.

Urine Interleukin-18 (IL-18) as a Biomarker of Total-Body Irradiation: A Preliminary Study in Nonhuman Primates.

We have reported that circulating IL-18 can be used as a radiation biomarker in mice, minipigs and nonhuman primates (NHPs, Macaca mulatta). Here, we report the levels of IL-18 in individual NHP's urine before and at 6 h-7 days after 5.0, 6.5 and 8.5 Gy 60Co total-body irradiation (TBI) using enzyme linked immunosorbent assay (ELISA). Six animals (3.5-5.5 kg, 3-4 years old) per radiation dose were investigated. Correlation values between urine IL-18 and blood cell counts and serum chemistry parameters including lactate dehydrogenase (LDH), lipase, and serum total protein (TP), as well as between urine IL-18 and 60-day survival, were analyzed. Our data, to the best of our knowledge, for the first time, demonstrate that concentrations of urine IL-18 from irradiated NHPs were increased in a radiation dose-dependent manner compared to pre-TBI levels in samples from these animal (N = 18, 11.02 ± 1.3 pg/ml). A 5.0 Gy low dose of radiation (∼LD10/60) did not increase urine IL-18 levels. In contrast, high-dose TBI significantly increased urine IL-18 at day 1 to day 5 in a bell-shaped time course, reaching a peak of 5- to 10-fold of control levels on day 3 after 6.5 Gy (∼LD50/60) and 8.5 Gy (∼LD90/60), respectively. Statistical analysis using receiver operator characteristic (ROC) and MultiROC analysis indicated that white blood cell and platelet counts, serum LDH, lipase and TP, when combined with urine IL-18, provide discriminatory predictors of total-body radiation injury with a very high ROC area of 0.98. Urine IL-18 measurement, as an early prognostic indicator of survival, may facilitate rapid detection of lethal doses of radiation, based on the currently available data set.

MODELING H-ARS USING HEMATOLOGICAL PARAMETERS: A COMPARISON BETWEEN THE NON-HUMAN PRIMATE AND MINIPIG.

Multiple hematological biomarkers (i.e. complete blood counts and serum chemistry parameters) were used in a multivariate linear-regression fit to create predictive algorithms for estimating the severity of hematopoietic acute radiation syndrome (H-ARS) using two different species (i.e. Göttingen Minipig and non-human primate (NHP) (Macacca mulatta)). Biomarker data were analyzed prior to irradiation and between 1-60 days (minipig) and 1-30 days (NHP) after irradiation exposures of 1.6-3.5 Gy (minipig) and 6.5 Gy (NHP) 60Co gamma ray doses at 0.5-0.6 Gy min-1 and 0.4 Gy min-1, respectively. Fitted radiation risk and injury categorization (RRIC) values and RRIC prediction percent accuracies were compared between the two models. Both models estimated H-ARS severity with over 80% overall predictive power and with receiver operating characteristic curve area values of 0.884 and 0.825. These results based on two animal radiation models support the concept for the use of a hematopoietic-based algorithm for predicting the risk of H-ARS in humans.

Is lymphocyte adenosine a diagnostic marker of clinical malignant hyperthermia? A pilot study.

OBJECTIVE: Malignant hyperthermia is a pharmacogenetic disorder typically triggered by potent inhalation anesthetics and/or the depolarizing muscle relaxant succinylcholine in malignant hyperthermia-susceptible individuals. Since lymphocytes express the same Ca channel mutation found in malignant hyperthermia-susceptible muscle, we investigated agonist-induced adenosine formation in lymphocytes as an index of sarcoplasmic reticulum Ca-release-induced adenosine 5'-triphosphate turnover as a potential minimally invasive functional malignant hyperthermia assay. DESIGN: Application of lymphocytes for malignant hyperthermia diagnosis. SETTING: Hospitals and university laboratory. SUBJECTS: Malignant hyperthermia-susceptible patients (n = 13) and normal subjects (n = 11). INTERVENTIONS: Adenosine formation due to malignant hyperthermia-triggering agent halothane or the ryanodine receptor Ca channels agonist 4-chloro-m-cresol was compared in blood lymphocytes from malignant hyperthermia-susceptible patients and normal subjects. MEASUREMENTS AND MAIN RESULTS: Cai and adenosine were measured in fresh or immortalized blood lymphocytes incubated with 0-10 mM 4-chloro-m-cresol or 0-10.7 mM halothane. Cai levels were significantly higher in immortalized malignant hyperthermia-susceptible B cells treated with 0.75 mM 4-chloro-m-cresol relative to controls. Similarly, at 1 mM 4-chloro-m-cresol or 0.96 mM halothane, adenosine levels were significantly higher in malignant hyperthermia-susceptible lymphocytes or immortalized B cells relative to controls. Receiver-operating characteristic analyses showed areas under the 4-chloro-m-cresol receiver-operating characteristic curves near more than or equal to 0.96 (p ≈ 0.0001), suggesting that 4-chloro-m-cresol-induced adenosine could readily distinguish between malignant hyperthermia-susceptible and normal controls cells. CONCLUSIONS: Both 4-chloro-m-cresol and halothane caused adenosine accumulation in blood lymphocytes. Adenosine accumulation was markedly increased in malignant hyperthermia-susceptible lymphocytes compared with controls reflecting higher than normal adenosine 5'-triphosphate degradation in the malignant hyperthermia-susceptible cells. Although 4-chloro-m-cresol receiver-operating characteristic curves revealed that adenosine accumulation could readily distinguish between normal and malignant hyperthermia-susceptible lymphocytes, independent confirmation is required with a substantially larger number of enrolled subjects to correctly appreciate the clinical utility of the novel lymphocyte-adenosine protocol for malignant hyperthermia testing.

Acetate transiently inhibits myocardial contraction by increasing mitochondrial calcium uptake.

BACKGROUND: There is a close relationship between cardiovascular disease and cardiac energy metabolism, and we have previously demonstrated that palmitate inhibits myocyte contraction by increasing Kv channel activity and decreasing the action potential duration. Glucose and long chain fatty acids are the major fuel sources supporting cardiac function; however, cardiac myocytes can utilize a variety of substrates for energy generation, and previous studies demonstrate the acetate is rapidly taken up and oxidized by the heart. In this study, we tested the effects of acetate on contractile function of isolated mouse ventricular myocytes. RESULTS: Acute exposure of myocytes to 10 mM sodium acetate caused a marked, but transient, decrease in systolic sarcomere shortening (1.49 ± 0.20% vs. 5.58 ± 0.49% in control), accompanied by a significant increase in diastolic sarcomere length (1.81 ± 0.01 µm vs. 1.77 ± 0.01 µm in control), with a near linear dose response in the 1-10 mM range. Unlike palmitate, acetate caused no change in action potential duration; however, acetate markedly increased mitochondrial Ca(2+) uptake. Moreover, pretreatment of cells with the mitochondrial Ca(2+) uptake blocker, Ru-360 (10 µM), markedly suppressed the effect of acetate on contraction. CONCLUSIONS: Lehninger and others have previously demonstrated that the anions of weak aliphatic acids such as acetate stimulate Ca(2+) uptake in isolated mitochondria. Here we show that this effect of acetate appears to extend to isolated cardiac myocytes where it transiently modulates cell contraction.

Application of multivariate modeling for radiation injury assessment: a proof of concept.

Multivariate radiation injury estimation algorithms were formulated for estimating severe hematopoietic acute radiation syndrome (H-ARS) injury (i.e., response category three or RC3) in a rhesus monkey total-body irradiation (TBI) model. Classical CBC and serum chemistry blood parameters were examined prior to irradiation (d 0) and on d 7, 10, 14, 21, and 25 after irradiation involving 24 nonhuman primates (NHP) (Macaca mulatta) given 6.5-Gy (60)Co Υ-rays (0.4 Gy min(-1)) TBI. A correlation matrix was formulated with the RC3 severity level designated as the "dependent variable" and independent variables down selected based on their radioresponsiveness and relatively low multicollinearity using stepwise-linear regression analyses. Final candidate independent variables included CBC counts (absolute number of neutrophils, lymphocytes, and platelets) in formulating the "CBC" RC3 estimation algorithm. Additionally, the formulation of a diagnostic CBC and serum chemistry "CBC-SCHEM" RC3 algorithm expanded upon the CBC algorithm model with the addition of hematocrit and the serum enzyme levels of aspartate aminotransferase, creatine kinase, and lactate dehydrogenase. Both algorithms estimated RC3 with over 90% predictive power. Only the CBC-SCHEM RC3 algorithm, however, met the critical three assumptions of linear least squares demonstrating slightly greater precision for radiation injury estimation, but with significantly decreased prediction error indicating increased statistical robustness.

A porcine model of complement-mediated infusion reactions to drug carrier nanosystems and other medicines.

Intravenous administration of low (milligram) doses of nanoparticulate materials in pigs can lead to acute cardiopulmonary, hemodynamic, hematological, biochemical and dermatological changes within minutes, mimicking the human infusion (or anaphylactoid) reactions to many state-of-the-art (nano)medicines and biologicals. Because of the causal role of complement (C) activation, the phenomenon was called C activation-related pseudoallergy (CARPA). This review summarizes the available information on porcine CARPA caused by different liposomes and polymers. It provides methodical details of the model and addresses the quantitation, sensitivity, specificity, reproducibility and variability of symptoms caused by different reactogenic drugs. We describe a unique feature of the model: the rise of tachyphylaxis (self-induced tolerance) as a function of structural properties of reactogenic agents. For drugs that cause tachyphylactic CARPA, such as liposomal doxorubicin (Doxil), the review recapitulates a recently reported method of desensitization, which may prevent this, as well as many similar hypersensitivity reactions. In explaining the underlying mechanism of tachyphylactic CARPA, a new theory on "double hit" is outlined, wherein the pulmonary intravascular macrophages (PIM cells) of pigs give aggravated response to simultaneous stimulation of their anaphylatoxin and other surface receptors (e.g., toll-like, PAMP, DAMP or mannose) that recognize vesicle surface molecular patterns. The porcine CARPA model might provide unique advantages in studying the mechanism of severe hypersensitivity reactions in man to i.v. drugs, as well as in identifying drugs and drug carriers that may cause such reactions.

Prevention of infusion reactions to PEGylated liposomal doxorubicin via tachyphylaxis induction by placebo vesicles: a porcine model.

PEGylated liposomal doxorubicin (Doxil) has been used in cancer chemotherapy for 16 years. Clinical experience shows that it can cause mild-to-severe hypersensitivity (infusion) reactions, which are manifestations of complement (C) activation-related pseudoallergy (CARPA). Although in most cases CARPA is inconsequential, a main symptom, cardiopulmonary distress, may be life threatening in hypersensitive individuals. To date, the prevention of Doxil-induced CARPA is based on premedication and a slow infusion protocol. The present study suggests desensitization by Doxil-like empty liposomes, called placebo Doxil (Doxebo), as an alternative strategy, which is based on the tachyphylactic nature of Doxil reactions. Doxebo-induced tolerance to Doxil was shown to develop within minutes and to be specific to Doxil-like PEGylated liposomes. The procedure of desensitization involves slow, low-dose pre-infusion of Doxebo before Doxil treatment which minimizes the ensuing physiological changes or keeps them subclinical. Although the mechanism of tolerance induction is not yet clear, the effector arm of C response is unlikely to be affected, as the vascular reactivity of desensitized pigs to zymosan remains intact. Desensitization with empty vesicles represents a novel approach for reducing the risk of anaphylactic reactions to drug carrier liposomes. The underlying immediate, most likely passive silencing of an innate immune response may represent a novel mechanism of tolerance induction which may work for other reactogenic nanosystems as well.

Case report highlighting how wound path identification on CT can help identify organ damage in abdominal blast injuries.

Penetrating trauma is frequently encountered in forward deployed military combat hospitals. Abdominal blast injuries represent nearly 11% of combat injuries, and multiplanar computed tomography imaging is optimal for injury assessment and surgical planning. We describe a multiplanar approach to assessment of blast and ballistic injuries, which allows for more expeditious detection of missile tracts and damage caused along the path. Precise delineation of the trajectory path and localization of retained fragments enables time-saving and detailed evaluation of associated tissue and vascular injury. For consistent and reproducible documentation of fragment locations in the body, we propose a localization scheme based on Cartesian coordinates to report 3-dimensional locations of fragments and demonstrating the application in three cases of abdominal blast injury.

Liposome-induced complement activation and related cardiopulmonary distress in pigs: factors promoting reactogenicity of Doxil and AmBisome.

Hypersensitivity reactions to liposomal drugs, often observed with Doxil and AmBisome, can arise from activation of the complement (C) system by phospholipid bilayers. To understand the mechanism of this adverse immune reaction called C activation-related pseudoallergy (CARPA), we analyzed the relationship among liposome features, C activation in human serum in vitro, and liposome-induced cardiovascular distress in pigs, a model for human CARPA. Among the structural variables (surface charge, presence of saturated, unsaturated, and PEGylated phospholipids, and cisplatin vs. doxorubicin inside liposomes), high negative surface charge and the presence of doxorubicin were significant contributors to reactogenicity both in vitro and in vivo. Morphological analysis suggested that the effect of doxorubicin might be indirect, via distorting the sphericity of liposomes and, if leaked, causing aggregation. The parallelism among C activation, cardiopulmonary reactions in pigs, and high rate of hypersensitivity reactions to Doxil and AmBisome in humans strengthens the utility of the applied tests in predicting the risk of CARPA. FROM THE CLINICAL EDITOR: The authors studied complement activation-related pseudoallergy (CARPA) in a porcine model and demonstrate that high negative surface charge and drug effects leading to distortion of liposome sphericity might be the most critical factors leading to CARPA. The applied tests might be used to predict CARPA in humans.

Cartesian positioning system for localization of blast and ballistic fragments: a phantom-based pilot study.

Our purpose was to demonstrate the consistency of radiologists' three-dimensional measurements of simulated blast fragment locations in vitro in an effort to objectively localize retained fragments and wound paths. We designed a phantom consisting of 10 nail heads (simulating blast fragments) glued to wooden pegs that were randomly situated at distances from a reference point within a plastic tub. The x, y, and z coordinates of simulated fragments were recorded in Cartesian 3-space relative to the reference point. Computed tomography images of the phantom were acquired. Differences in x, y, and z positions as determined by three observers were summed for each fragment. Agreement between recordings of coordinates across readers was assessed using the intraclass correlation coefficient. Summed differences in coordinate positions as determined by readers ranged between 0.00 and 1.204 cm (mean: 0.732 cm). Across readers, the intraclass correlation coefficient for each dimension was >0.99. We found excellent agreement among readers with minimal discrepancy of measured locations of simulated fragments. Our results provide a foundation for trajectory analysis necessary to lead to automated organ damage reporting for immediate assessment in the emergency department and for forensic investigation and long-term epidemiological analysis.

Parahydrogen-induced polarization (PHIP) hyperpolarized MR receptor imaging in vivo: a pilot study of 13C imaging of atheroma in mice.

MR techniques using hyperpolarized (13)C have successfully produced examples of angiography and intermediary metabolic imaging, but, to date, no receptor imaging has been attempted. The goal of this study was to synthesize and evaluate a novel hyperpolarizable molecule, 2,2,3,3-tetrafluoropropyl 1-(13)C-propionate-d(2,3,3) (TFPP), for the detection of atheromatous plaques in vivo. TFPP binds to lipid bilayers and its use in hyperpolarized MR could prove to be a major step towards receptor imaging. The precursor, 2,2,3,3-tetrafluoropropyl 1-(13)C-acrylate-d(2,3,3) (TFPA), binds to 1,2-dimyristoylphosphatidylcholine lipid bilayers with a 1.6-ppm chemical shift in the (19)F MR spectrum. This molecule was designed to be hyperpolarized through the addition of parahydrogen to the (13)C-acrylate moiety by parahydrogen-induced polarization. TFPA was hyperpolarized to TFPP to an extent similar to that of the hydroxyethylacrylate to hydroxyethylpropionate transition: 17 ± 4% for TFPP versus 20% for hydroxyethylpropionate; T(1) relaxation times (45 ± 2 s versus 55 ± 2 s) were comparable and the hyperpolarized properties of TFPP were characterized. Hydroxyethylacrylate, like TFPA, has a chemical structure with an acrylate moiety, but does not contain the lipid-binding tetrafluoropropyl functional group. Hyperpolarized TFPP binds to the lipid bilayer, appearing as a second, chemically shifted (13)C hyperpolarized MR signal with a further reduction in the longitudinal relaxation time (T(1) = 21 ± 1 s). In aortas harvested from low-density lipoprotein receptor knock-out mice fed with a high-fat diet for 9 months, and in which atheroma is deposited in the aorta and heart, TFPP showed greater binding to lipid on the intimal surface than in control mice fed a normal diet. When TFPP was hyperpolarized and administered in vivo to atheromatous mice in a pilot study, increased binding was observed on the endocardial surface of the intact heart compared with normally fed controls. Hyperpolarized TFPP has bio-sensing specificity for lipid, coupled with a 42,000-fold sensitivity gain in the MR signal at 4.7 T. Binding of TFPP with lipids results in the formation of a characteristic second peak in MRS. TFPP therefore has the potential to act as an in vivo molecular probe for atheromatous plaque imaging and may serve as a model of receptor-targeted bio-imaging with enhanced MR sensitivity.

CT-based ballistic wound path identification and trajectory analysis in anatomic ballistic phantoms.

PURPOSE: To evaluate the accuracy of computed tomography (CT)-based ballistic wound path identification in phantoms by determining the agreement between actual shooting angles and both trajectory angles measured with a picture archiving and communication system (PACS) angle tool and angles calculated from x, y, z coordinates of the entrance and exit points. MATERIALS AND METHODS: In this institutional review board-approved model study, two simulated legs were shot by a trained marksman from 50 yards at six clinometer-measured angles with a 0.30-06 rifle and then scanned at multidetector CT. Radiologists measured the wound path angles on paracoronal reformations by using a PACS angle tool. Observers determined the Cartesian coordinates of the entrance and exit points of the wound paths on axial CT images by using detailed instructions. Angles were calculated from these coordinates by using a computer arctangent function. Agreement between the angles was evaluated with Bland-Altman plots. Means, ranges, and standard deviations of the angles also were determined. RESULTS: Radiologists identified all six wound paths on the CT images. The PACS tool-based measured and coordinate-based calculated angles were within 5° of the shooting angles. Results indicated that in larger study populations, one can be 91% confident that future coordinate-based angle calculations will differ from the actual shooting angle by no more than 5° and 95% confident that PACS tool-based angle measurements will differ from the actual shooting angles by no more than 4.5°. One can be 95% confident that future coordinate-based angle calculations will differ from PACS angle measurements by no more than 4.02°. CONCLUSION: Study results demonstrated the feasibility of consistent wound path identification and the accuracy of trajectory angle determination in models with use of multidetector CT.

Lymphocyte-based determination of susceptibility to malignant hyperthermia: a pilot study in swine.

BACKGROUND: Malignant hyperthermia susceptibility (MHS) is diagnosed by an invasive in vitro caffeine-halothane contracture test (CHCT) carried out on biopsied skeletal muscle tissue. We are presenting a novel blood test approach for malignant hyperthermia testing in a swine model. Our main aim was to determine whether adenosine production from lymphocytes after 4-chloro-m-cresol (4CmC) stimulation distinguishes homozygous swine carrying the Arg615Cys mutation in the ryanodine receptor type 1 (RyR1) gene (MHS swine) from normal swine. METHODS: Lymphocytes were isolated from arterial blood (40 ml) obtained from MHS (n = 7) and normal (n = 7) swine. Cells were suspended in Hank's balanced salt solution and treated with 4CmC (0-10 mm) at 37°C in the presence of adenosine deaminase inhibitor. After termination and purification of samples, aliquots (50 µl) were assayed for adenosine content using high performance liquid chromatography. RESULTS: Baseline adenosine levels before stimulating lymphocytes with 4CmC were 0.025 ± 0.004 and 0.041 ± 0.006 µm (mean ± SEM) in lymphocytes from normal and MHS swine, respectively (P = 0.125). Maximum responses were achieved at 1 mm 4CmC for both cell-line groups. Adenosine levels after stimulation with 4CmC (1 mm) were 0.185 ± 0.009 and 0.397 ± 0.038 µm in lymphocytes from normal and MHS swine, respectively (P = 0.0035). There was no overlap between adenosine levels in stimulated lymphocytes from MHS and normal swine. CONCLUSION: 4CmC stimulation of porcine lymphocytes induces increased adenosine formation in MHS cells relative to those from normal swine; evaluation of adenosine formation in response to RyR1 agonists in human lymphocytes is needed.

Cardiovascular applications of hyperpolarized contrast media and metabolic tracers.

Modern hyperpolarization technology enhances the recordable magnetic resonance signal four to five orders of magnitude, making in vivo assessments of tracer pathways and metabolic compartments feasible. Existing hyperpolarization instrumentation and previous tracer studies using hydroxyethylpropionate (HEP) as an extracellular marker and 14-carbon label pyruvate as examples are described and reviewed as applicable to the working heart. Future metabolic imaging based on the use of hyperpolarized pyruvate needs to consider extra- and intra-cellular label dilution due to glycolysis, lactate oxidation and protein degradation. This dilution can substantially decrease the recordable signals from PDH flux (oxidative decarboxylation of pyruvate) and other pyruvate pathways. The review of previous literature and data suggests that the (13)C-alanine signal is a better index of mitochondrially oxidized pyruvate than L-lactate. These facts and considerations will help in the interpretation of the in vivo recorded hyperpolarization signals of metabolic tracers and contrast media.