Arterial Plasma Samples Research Articles

Soluble Glycoprotein VI Levels Assessed Locally within the Extra- and Intracerebral Circulation in Hyper-Acute Thromboembolic Stroke: A Pilot Study.

Background: Severe acute ischemic stroke (AIS) is mainly caused by thromboembolism originating from symptomatic carotid artery (ICA) stenosis or in the heart due to atrial fibrillation. Glycoprotein VI (GPVI), a principal platelet receptor, facilitates platelet adherence and thrombus formation at sites of vascular injury such as symptomatic ICA stenosis. The shedding of GPVI from the platelet surface releases soluble GPVI (sGPVI) into the circulation. Here, we aimed to determine whether sGPVI can serve as a local biomarker to differentiate between local atherosclerotic and systemic cardiac thromboembolism in AIS. Methods: We conducted a cohort study involving 105 patients undergoing emergency endovascular thrombectomy (EVT) for anterior circulation stroke. First, sGPVI concentrations were measured in systemic arterial plasma samples collected at the ipsilateral ICA level, including groups with significantly (≥50%) stenotic and non-stenotic arteries. A second sample, taken from the intracerebral pial circulation, was used to assess GPVI shedding locally within the ischemic brain. Results: Our analysis revealed no significant increase in systemic sGPVI levels in patients with symptomatic ≥ 50% ICA stenosis (3.2 [95% CI 1.5-5.0] ng/mL; n = 33) compared with stroke patients without significant ICA stenosis (3.2 [95% CI 2.3-4.2] ng/mL; n = 72). Additionally, pial blood samples, reflecting intravascular molecular conditions during collateral flow, showed similar sGPVI levels when compared to the systemic ICA samples in both groups. Conclusions: Our findings indicate that GPVI is not locally cleaved and shed into the bloodstream in significant amounts during hyper-acute ischemic stroke, neither at the level of symptomatic ICA nor intracranially during collateral blood supply. Therefore, sGPVI does not appear to be suitable as a local stroke biomarker despite strong evidence of a major role for GPVI-signaling in stroke pathophysiology.

Beta2-adrenergic agonist salbutamol exhibits enantioselective disposition in skeletal muscle of lean young men following oral administration.

Salbutamol is a common short-acting beta2-adrenergic agonist used in treatment of asthma and exercise-induced bronchoconstriction but also possesses anabolic and metabolic actions in skeletal muscle. As a chiral compound, salbutamol is a racemic 1:1 mixture of two enantiomers, (R)-salbutamol and (S)-salbutamol, which exhibit divergent pharmacokinetic and pharmacodynamic actions. Despite salbutamol being available for decades, information on the enantioselective disposition of salbutamol enantiomers in human skeletal muscle is absent. In this study, we determined concentrations of (R)-salbutamol and (S)-salbutamol by UHPLC-MS/MS in arterial plasma and vastus lateralis muscle samples from 12 lean young men 2½ and 7 h following ingestion of 24 mg oral salbutamol. Mean (range) arterial plasma concentrations were 10-fold higher (p < 0.001) for (S)-salbutamol than (R)-salbutamol, being 33(9-62) and 49(30-84) ng·mL-1 for (S)-salbutamol and 4 (1-6) and 4 (2-5) ng·mL-1 for (R)-salbutamol 2½ and 7 h following administration, respectively, reflecting faster elimination of the (R)-enantiomer. Mean (range) muscle concentrations were higher (p < 0.001) for (S)-salbutamol than (R)-salbutamol 2½h (0.17 [0.1-0.26] vs. 0.04 [0.02-0.06]) and 7 h (0.31 [0.21-0.46] vs. 0.06 [0.04-0.12] ng·mgd.w. -1) after administration. However, muscle:plasma partition coefficient was two-fold higher (p < 0.001) for (R)-salbutamol than (S)-salbutamol 7 h following administration. These observations demonstrate that oral salbutamol exhibits enantioselective disposition in systemic circulation and muscle favoring the (S)-enantiomer but with higher relative partitioning of the (R)-enantiomer in skeletal muscle. Furthermore, the concentration-time profiles of salbutamol enantiomers are different in skeletal muscle and systemic circulation following oral ingestion. These findings have implications for the application of chiral switch (R)-salbutamol in doping control.

Effect of albumin substitution on pharmacokinetics of piperacillin/tazobactam in patients with severe burn injury admitted to the ICU.

Pathophysiological changes in severely burned patients alter the pharmacokinetics (PK) of anti-infective agents, potentially leading to subtherapeutic concentrations at the target site. Albumin supplementation, to support fluid resuscitation, may affect pharmacokinetic properties by binding drugs. This study aimed to investigate the PK of piperacillin/tazobactam in burn patients admitted to the ICU before and after albumin substitution as total and unbound concentrations in plasma. Patients admitted to the ICU and scheduled for 4.5 g piperacillin/tazobactam administration and 200 mL of 20% albumin substitution as part of clinical routine were included. Patients underwent IV microdialysis, and simultaneous arterial plasma sampling, at baseline and multiple timepoints after drug administration. PK analysis of total and unbound drug concentrations under steady-state conditions was performed before and after albumin supplementation. A total of seven patients with second- to third-degree burns involving 20%-60% of the total body surface were enrolled. Mean (SD) AUC0-8 (h·mg/L) of total piperacillin/tazobactam before and after albumin substitution were 402.1 (242)/53.2 (27) and 521.8 (363)/59.7 (32), respectively. Unbound mean AUC0-8 before and after albumin supplementation were 398.9 (204)/54.5 (25) and 456.4 (439)/64.5 (82), respectively. Albumin supplementation had little impact on the PK of piperacillin/tazobactam. After albumin supplementation, there was a numerical increase in mean AUC0-8 of total and unbound piperacillin/tazobactam, whereas similar Cmax values were observed. Future studies may investigate the effect of albumin supplementation on drugs with a higher plasma protein binding.

Parametric and non-parametric Poisson regression for modelling of the arterial input function in positron emission tomography

Full quantification of Positron Emission Tomography (PET) requires an arterial input function (AIF) for measurement of certain targets, or using particular radiotracers, or for the quantification of specific outcome measures. The AIF represents the measurement of radiotracer concentrations in the arterial blood plasma over the course of the PET examination. Measurement of the AIF is prone to error as it is a composite measure created from the combination of multiple measurements of different samples with different equipment, each of which can be sources of measurement error. Moreover, its measurement requires a high degree of temporal granularity for early time points, which necessitates a compromise between quality and quantity of recorded samples. For these reasons, it is often desirable to fit models to this data in order to improve its quality before using it for quantification of radiotracer binding in the tissue. The raw observations of radioactivity in arterial blood and plasma samples are derived from radioactive decay, which is measured as a number of recorded counts. Count data have several specific properties, including the fact that they cannot be negative as well as a particular mean-variance relationship. Poisson regression is the most principled modelling strategy for working with count data, as it both incorporates and exploits these properties. However, no previous studies to our knowledge have taken this approach, despite the advantages of greater efficiency and accuracy which result from using the appropriate distributional assumptions. Here, we implement a Poisson regression modelling approach for the AIF as proof-of-concept of its application. We applied both parametric and non-parametric models for the input function curve. We show that a negative binomial distribution is a more appropriate error distribution for handling overdispersion. Furthermore, we extend this approach to a hierarchical non-parametric model which is shown to be highly resilient to missing data. We thus demonstrate that Poisson regression is both feasible and effective when applied to AIF data, and propose that this is a promising strategy for modelling blood count data for PET in future.

Plasma Metabolome Alterations Discriminate between COVID-19 and Non-COVID-19 Pneumonia.

Pneumonia is a common cause of morbidity and mortality and is most often caused by bacterial pathogens. COVID-19 is characterized by lung infection with potential progressive organ failure. The systemic consequences of both disease on the systemic blood metabolome are not fully understood. The aim of this study was to compare the blood metabolome of both diseases and we hypothesize that plasma metabolomics may help to identify the systemic effects of these diseases. Therefore, we profiled the plasma metabolome of 43 cases of COVID-19 pneumonia, 23 cases of non-COVID-19 pneumonia, and 26 controls using a non-targeted approach. Metabolic alterations differentiating the three groups were detected, with specific metabolic changes distinguishing the two types of pneumonia groups. A comparison of venous and arterial blood plasma samples from the same subjects revealed the distinct metabolic effects of pulmonary pneumonia. In addition, a machine learning signature of four metabolites was predictive of the disease outcome of COVID-19 subjects with an area under the curve (AUC) of 86 ± 10 %. Overall, the results of this study uncover systemic metabolic changes that could be linked to the etiology of COVID-19 pneumonia and non-COVID-19 pneumonia.

RF12 | PSUN106 Quantifying Liver and Brain Levels of 11β-hydroxysteroid Dehydrogenase Type 1 in Obesity Using Positron Emission Tomography Imaging

Abstract Objectives Cortisol is known to promote adipocyte differentiation and maturation, and prolonged exposure to excess cortisol contributes to the development of obesity and metabolic dysregulation. The intracellular 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) enzyme catalyzes the conversion of inactive cortisone to active cortisol. Recently, we demonstrated reduced brain 11β-HSD1 levels in vivo with increasing body mass index (BMI), using positron emission tomography (PET) radioligands [11C]- and [18F]AS2471907(1). PET imaging can simultaneously quantify uptake in multiple organs (e.g., liver/brain). [18F]AS2471907 has high uptake and specific binding in the brain and liver(2). Liver and brain 11β-HSD1 measured by PET imaging may help elucidate the roles of cortisol activation in the setting of metabolic dysregulation. Thus, we performed PET imaging studies to examine 11β-HSD1 levels in the liver and brain. Methods Nine individuals (5F/4M) with a range of BMIs (22.6-34.4 kg/m2) underwent a 90 min PET/CT acquisition with arterial plasma sampling after injection of [18F]AS2471907. Regions-of-interest (ROI) for the liver were manually drawn on a summed PET image (60-90 min). Seventeen brain ROIs were selected from the anatomical automatic labeling (AAL) template and applied to the dynamic PET images to generate time-activity-curves (TACs). Brain volume of distribution (VT, mL/cm3), the target tissue to plasma ratio of radioligand at equilibrium, was estimated for each ROI using the multilinear analysis-1 method with plasma input function. Mean whole-brain VT values were calculated by averaging all ROIs. Given the appearance of irreversible kinetics in the liver, Ki (min-1), the rate of irreversible tracer uptake, was calculated using the Patlak method. Results Qualitative brain and liver uptake was assessed by examining a summed PET image (SUV 60-90min). The parent fraction in plasma was 88±1% at 90 min, indicating negligible plasma radiolabeled-metabolites contributing the measured signal in the liver. Kinetic modeling estimates demonstrated decreasing whole brain VT with increasing BMI (R2=0.53), similar to our previously published study examining only brain in a larger cohort(1). Patlak methods provided good estimates of Ki. In an opposite manner from the brain, correlations seen in the liver using Ki were positively correlated with BMI (R2=0.57). Conclusions These preliminary studies suggest obesity is associated with increased 11β-HSD1 levels in the liver but decreased 11β-HSD1 levels in the brain. Clinically, liver specific 11β-HSD1 inhibitors may prove beneficial in treating metabolic sequalae of obesity such non-alcoholic fatty liver disease (NAFLD) by inhibiting activity of higher 11β-HSD1 levels. Further studies in NAFLD and obesity are necessary to determine appropriate populations that would benefit from 11β-HSD1 inhibitors and longitudinal in vivo PET imaging to help inform development of future therapeutics.

Insulin Infusion Is Linked to Increased NPPC Expression in Muscle and Plasma C-type Natriuretic Peptide in Male Dogs.

The purpose of this study was to assess insulin-stimulated gene expression in canine skeletal muscle with a particular focus on NPPC, the gene that encodes C-type natriuretic peptide, a key hormonal regulator of cardiometabolic function. Four conscious canines underwent hyperinsulinemic, euglycemic clamp studies. Skeletal muscle biopsy and arterial plasma samples were collected under basal and insulin-stimulated conditions. Bulk RNA sequencing of muscle tissue was performed to identify differentially expressed genes between these 2 steady-state conditions. Our results showed that NPPC was the most highly expressed gene in skeletal muscle in response to insulin infusion, rising 4-fold between basal and insulin-stimulated conditions. In support of our RNA sequencing data, we found that raising the plasma insulin concentration 15-fold above basal elicited a 2-fold (P = 0.0001) increase in arterial plasma concentrations of N-terminal prohormone C-type natriuretic peptide. Our data suggest that insulin may play a role in stimulating secretion of C-type natriuretic peptide by skeletal muscle. In this context, C-type natriuretic peptide may act in a paracrine manner to facilitate muscle–vascular bed crosstalk and potentiate insulin-mediated vasodilation. This could serve to enhance insulin and glucose delivery, particularly in the postprandial absorptive state.

Antimicrobial REG3A and Network Inflammatory Proteins Reflect Stroke Severity and Functional Impairment During Ischemic Stroke

Introduction Regenerating Family Member 3 Alpha (REG3A) is an antimicrobial protein secreted by the intestine and pancreas and is involved in immune-mediated inflammatory responses. In healthy individuals, REG3A is minimally expressed. However, acute-phase inflammatory mediators such as IL6 upregulate REG3A during inflammation, trauma, and cell dysregulation. The University of Kentucky Blood and Clot Thrombectomy Registry and Collaboration (BACTRAC) protocol utilizes thrombectomy to isolate intracranial arterial blood (i.e. distal to thrombus) and systemic arterial blood (i.e. carotid) to better understand the pathophysiology of stroke. Initial proteomic analyses of 184 plasma proteins sampled during thrombectomy found that systemic arterial REG3A was increased during large vessel occlusion. The function of REG3A in neuroinflammation sequela of ischemic stroke has yet to be elucidated. Here, we examined the association of plasma REG3A levels with other signaling proteins in systemic plasma during stroke, and its correlation with clinical outcomes of stroke. Methods Systemic arterial plasma samples from 35 thrombectomy subjects underwent Proximity Extension Assay via Olink Proteomics. REG3A levels and inflammatory markers were examined using bivariate regressions. Biserial correlation examined the relationship between the National Institutes of Health Stroke Scale (NIHSS) upon hospital admission and REG3A. Multilinear regression determined predictability of infarct volume and edema volume by REG3A. Results: Systemic arterial levels of IL6 and REG3A were positively correlated (R=0.371, P=0.031). Higher levels of REG3A correlated with increased infarct and edema volume (R=0.416, P=0.014; R=0.418, P=0.014). Multilinear regression predicting infarct (R2=0.985,P<0.001) and edema volume (R2=0.998, P<0.001) each yielded a model with similar shared proteins, REG3A, CXCL11, TGFβ, IL17A, and NCAM1. NIHSS score at admission positively correlated with REG3A( R= 0.373, P=0.032). Conclusions Within a network of proteins, REG3A was predictive of infarct severity and increased neurologic deficits in patients with ELVO. Results of IL6 and REG3A uphold similar associations reported within the literature. Models predicting stroke severity yielded potential protein partners with REG3A, such as IL17, CXCL11, and TGFβ, which interact with REG3A in other inflammatory processes. REG3A function in stroke has yet to be understood, however this study reports novel findings demonstrating the relationship between REG3A and outcomes of stroke. Examining these associations will be critical in defining REG3A function during neuroinflammation and as a potential biomarker of large vessel occlusive stroke predicting clinical outcomes.

11C-PK11195 plasma metabolization has the same rate in multiple sclerosis patients and healthy controls: a cross-sectional study.

11C-PK11195 is a positron emitter tracer used for Positron Emission Tomography (PET) imaging of innate immune cell activation in studies of neuroinflammatory diseases. For the image quantitative analysis, it is necessary to quantify the intact fraction of this tracer in the arterial plasma during imaging acquisition (plasma intact fraction). Due to the complexity and costs involved in this analysis it is important to evaluate the real necessity of individual analysis in each 11C-PK11195 PET imaging acquisition. The purpose of this study is to compare 11C-PK11195 plasma metabolization rate between healthy controls and multiple sclerosis (MS) patients and evaluate the interference of sex, age, treatment, and disease phenotype in the tracer intact fraction measured in arterial plasma samples. 11C-PK11195 metabolization rate in arterial plasma was quantified by high performance liquid chromatography in samples from MS patients (n = 50) and healthy controls (n = 23) at 20, 45, and 60 minutes after 11C-PK11195 injection. Analyses were also stratified by sex, age, treatment type, and MS phenotype. The results showed no significant differences in the metabolization rate of healthy controls and MS patients, or in the stratified samples. In conclusion, 11C-PK11195 metabolization has the same rate in patients with MS and healthy controls, which is not affected by sex, age, treatment, and disease phenotype. Thus, these findings could contribute to exempting the necessity for tracer metabolization determination in all 11C-PK11195 PET imaging acquisition, by using a population metabolization rate average. The study procedures were approved by the Ethics Committee for Research Projects Analysis of the Hospital das Clinicas of the University of Sao Paulo Medical School (approval No. 624.065) on April 23, 2014.

Pharmacokinetics and pharmacodynamics of l-methadone in isoflurane-anaesthetized and mechanically ventilated ponies

ObjectiveTo evaluate the pharmacokinetics and selected pharmacodynamic effects of a commercially available l-methadone/fenpipramide combination administered to isoflurane anaesthetized ponies. Study designProspective single-group interventional study. AnimalsA group of six healthy adult research ponies (four mares, two geldings). MethodsPonies were sedated with intravenous (IV) detomidine (0.02 mg kg–1) and butorphanol (0.01 mg kg–1) for an unrelated study. Additional IV detomidine (0.004 mg kg–1) was administered 85 minutes later, followed by induction of anaesthesia using IV diazepam (0.05 mg kg–1) and ketamine (2.2 mg kg–1). Anaesthesia was maintained with isoflurane in oxygen. Baseline readings were taken after 15 minutes of stable isoflurane anaesthesia. l-Methadone (0.25 mg kg–1) with fenpipramide (0.0125 mg kg–1) was then administered IV. Selected cardiorespiratory variables were recorded every 10 minutes and compared to baseline using the Wilcoxon signed-rank test. Adverse events were recorded. Arterial plasma samples for analysis of plasma concentrations and pharmacokinetics of l-methadone were collected throughout anaesthesia at predetermined time points. Data are shown as mean ± standard deviation or median and interquartile range (p < 0.05). ResultsPlasma concentrations of l-methadone showed a rapid initial distribution phase followed by a slower elimination phase which is best described with a two-compartment model. The terminal half-life was 44.3 ± 18.0 minutes, volume of distribution 0.43 ± 0.12 L kg–1 and plasma clearance 7.77 ± 1.98 mL minute–1 kg–1. Mean arterial blood pressure increased from 85 (±16) at baseline to 100 (±26) 10 minutes after l-methadone/fenpipramide administration (p = 0.031). Heart rate remained constant. In two ponies fasciculations occurred at different time points after l-methadone administration. Conclusions and clinical relevanceAdministration of a l-methadone/fenpipramide combination to isoflurane anaesthetized ponies led to a transient increase in blood pressure without concurrent increases in heart rate. Pharmacokinetics of l-methadone were similar to those reported for conscious horses administered racemic methadone.

Pharmacokinetics of ketamine and its major metabolites norketamine, hydroxynorketamine, and dehydronorketamine: a model-based analysis

BackgroundRecent studies show activity of ketamine metabolites, such as hydroxynorketamine, in producing rapid relief of depression-related symptoms and analgesia. To improve our understanding of the pharmacokinetics of ketamine and metabolites norketamine, dehydronorketamine, and hydroxynorketamine, we developed a population pharmacokinetic model of ketamine and metabolites after i.v. administration of racemic ketamine and the S-isomer (esketamine). Pharmacokinetic data were derived from an RCT on the efficacy of sodium nitroprusside (SNP) in reducing the psychotomimetic side-effects of ketamine in human volunteers. MethodsThree increasing i.v. doses of esketamine and racemic ketamine were administered to 20 healthy volunteers, and arterial plasma samples were obtained for measurement of ketamine and metabolites. Subjects were randomised to receive esketamine/SNP, esketamine/placebo, racemic ketamine/SNP, and racemic ketamine/placebo on four separate occasions. The time–plasma concentration data of ketamine and metabolites were analysed using a population compartmental model approach. ResultsThe pharmacokinetics of ketamine and metabolites were adequately described by a seven-compartment model with two ketamine, norketamine, and hydroxynorketamine compartments and one dehydronorketamine compartment with metabolic compartments in-between ketamine and norketamine, and norketamine and dehydronorketamine main compartments. Significant differences were found between S- and R-ketamine enantiomer pharmacokinetics, with up to 50% lower clearances for the R-enantiomers, irrespective of formulation. Whilst SNP had a significant effect on ketamine clearances, simulations showed only minor effects of SNP on total ketamine pharmacokinetics. ConclusionsThe model is of adequate quality for use in future pharmacokinetic and pharmacodynamic studies into the efficacy and side-effects of ketamine and metabolites. Clinical trial registrationDutch Cochrane Center 5359.

Assessing metabolic properties of dairy cows fed low quality straws by integrative arterial and venous metabolomics.

ObjectiveThis study was conducted to reveal potential metabolic differences of dairy cows fed corn stover (CS) and rice straw (RS) instead of alfalfa hay (AH) as main forage source.MethodsThirty multiparous mid-late lactation Holstein dairy cows were selected and randomly assigned to three diets, AH, CS, or RS (n = 10). After 13 weeks of the feeding trial, coccygeal arterial and superficial epigastric venous plasma samples were collected before morning feeding for gas chromatography time-of-flight/mass spectrometry analyses.ResultsIn the artery, 8 and 13 metabolites were detected as differential metabolites between AH and CS, and between AH and RS, respectively. The relative abundance of phenylpropanoate (log2fold change [FC]) = 1.30, 1.09), panthenol (log2FC = 2.36, 2.20), threitol (log2FC = 1.00, 1.07), and 3,7,12-trihydroxycoprostane (log2FC = 0.79, 0.78) were greater in both CS and RS than in AH, and tyrosine (log2FC = −0.32), phenylalanine (log2FC = −0.30), and pyruvic acid (log2FC = −0.30) were lower in RS than in AH. In the vein, 1 and 7 metabolites were detected as differential metabolites between AH and CS, and between AH and RS, respectively. By comparing AH and RS, we found that metabolic pathways of phenylalanine, tyrosine, and tryptophan biosynthesis and phenylalanine metabolism were enriched by integrative artery and vein analysis. Furthermore, AH and RS, arterial phenylpropanoate and 4-hydroxyproline were positively, and phenylalanine was negatively correlated with milk urea nitrogen. Finally, in AH and CS, arterial panthenol was negatively correlated with feed efficiency.ConclusionArterial metabolic profiles changed more than those in the veins from animals on three forage diets, differing in amino acids. We found that phenylalanine, tyrosine, and tryptophan biosynthesis and phenylalanine metabolism were restricted when cows were fed low-quality cereal straw diets.

Lifelong Physical Activity Determines Vascular Function in Late Postmenopausal Women.

The study evaluated the role of lifelong physical activity for leg vascular function in postmenopausal women (61 ± 1 yr). The study design was cross-sectional with three different groups based on self-reported physical activity level with regard to intensity and volume over the past decade: inactive (n = 14), moderately active (n = 12), and very active (n = 15). Endothelial-dependent and smooth muscle-dependent leg vascular function were assessed by ultrasound Doppler measurements of the femoral artery during infusion of acetylcholine (Ach), the nitric oxide (NO) donor sodium nitroprusside and the prostacyclin analog epoprostenol. Thigh muscle biopsies, arterial and venous plasma samples were obtained for assessment of vasodilator systems. The very active group was found to have 76% greater responsiveness to Ach compared with the sedentary group accompanied by 200% higher prostacyclin synthesis during Ach infusion. Smooth muscle cell responsiveness to sodium nitroprusside and epoprostenol was not different between groups. The protein amount of endothelial NO synthase and endogenous antioxidant enzymes in muscle tissue was higher in the very active than the inactive group. The moderately active group had a similar endothelial and smooth muscle cell responsiveness as the inactive group. A secondary comparison with a smaller group (n = 5) of habitually active young (24 ± 2 yr) women indicated that smooth muscle cell responsiveness and endothelial responsiveness are affected by age per se. This study shows that leg vascular function and the potential to form prostacyclin and NO in late postmenopausal women, is influenced by the extent of lifelong physical activity.

Skeletal muscle amino acid uptake is lower and alanine production is greater in late gestation intrauterine growth-restricted fetal sheep hindlimb.

In a sheep model of intrauterine growth restriction (IUGR) produced from placental insufficiency, late gestation fetuses had smaller skeletal muscle mass, myofiber area, and slower muscle protein accretion rates compared with normally growing fetuses. We hypothesized that IUGR fetal muscle develops adaptations that divert amino acids (AAs) from protein accretion and activate pathways that conserve substrates for other organs. We placed hindlimb arterial and venous catheters into late gestation IUGR (n = 10) and control (CON, n = 8) fetal sheep and included an external iliac artery flow probe to measure hindlimb AA uptake rates. Arterial and venous plasma samples and biceps femoris muscle were analyzed by mass spectrometry-based metabolomics. IUGR fetuses had greater abundance of metabolites enriched within the alanine, aspartate, and glutamate metabolism pathway compared with CON. Net uptake rates of branched-chain AA (BCAA) were lower by 42%-73%, and muscle ammoniagenic AAs (alanine, glycine, and glutamine) were lower by 107%-158% in IUGR hindlimbs versus CON. AA uptake rates correlated with hindlimb weight; the smallest hindlimbs showed net release of ammoniagenic AAs. Gene expression levels indicated a decrease in BCAA catabolism in IUGR muscle. Plasma purines were lower and plasma uric acid was higher in IUGR versus CON, possibly a reflection of ATP conservation. We conclude that IUGR skeletal muscle has lower BCAA uptake and develops adaptations that divert AAs away from protein accretion into alternative pathways that sustain global energy production and nitrogen disposal in the form of ammoniagenic AAs for metabolism in other organs.

Development of a thin layer chromatography method for plasma correction of [18F]fluorocholine metabolites in positron emission tomography quantification studies in humans

IntroductionAfter its intravenous injection, [18F]fluorocholine is oxidized by choline-oxidase into its main plasma metabolite, [18F]fluorobetaine. If PET kinetic modeling quantification of [18F]fluorocholine uptake is intended, the plasma input time-activity-curve of the parent tracer must be obtained, i.e., the fraction of the total plasma radioactivity corresponding to the nonmetabolized [18F]fluorocholine at each time has to be known. Hence our aim was to develop an easy-routine Thin-Layer-Chromatography (TLC) method to separate and quantify the relative fractions of [18F]fluorocholine and [18F]fluorobetaine as a function of time during PET imaging in humans. MethodsFirst, we tested several combinations of solvents systems and layers to select the one showing the best resolution on non-radioactive standards. Thereafter, [18F]fluorobetaine was obtained through chemical oxidation of an [18F]fluorocholine sample at diferent incubation times and we applied the selected TLC-system to aliquots of this oxidation solution, both in a saline and in human deproteinized plasma matrices. The plates were detected by a radio-TLC-scanner. This TLC-system was finally applied to arterial plasma samples from 9 patients with high-grade-glioma undergoing brain PET imaging and a parent fraction curve was obtained in each of them. ResultsA TLC-system based on Silica-Gel-60//MeOH-NH3 was selected from the choline/betaine non-radioactive standards assay. Radiochromatograms of [18F]fluorocholine oxidation solution yielded two separated and well-defined peaks, Rf = 0,03 ([18F]fluorocholine) and Rf = 0.78 (18F]fluorobetaine) consistent with those observed on non-radioactive standards. During the oxidation, the [18F]fluorocholine radioactivity peak decreased progressively at several incubation times, while the other peak ([18F]fluorobetaine) increased accordingly. The mean values of the parent fraction of [18F]fluorocholine of the 9 patients studied (mean+/-SD) were 94% ± 6%, 58% ± 15%, 43% ± 10%, 39% ± 6% and 37% ± 6% at 2.8 min, 5.8 min, 8.8 min, 11.7 min and 14.7 min post-injection, respectively. ConclusionsWe have developed a TLC-system, easy to perform in a standard radiopharmacy unit, that enables the metabolite correction of arterial input function of [18F]fluorocholine in patients undergoing PET oncologic quantitative imaging.

Analysis of capillary microsamples obtained from a skin-prick to measure vancomycin concentrations as a valid alternative to conventional sampling: A bridging study

A bridging study is presented to investigate the applicability of measuring vancomycin concentrations obtained by finger-prick. A total of 25 paired plasma samples, collected from finger prick as capillary microsampling and arterial plasma samples collected from an indwelling cannula as conventional sampling, were obtained from critically ill patients receiving vancomycin. The maximum concentration (Cmax) and the minimum concentration (Cmin) measured were 66.2 mg/L and 29.7 mg/L for capillary microsampling and 78.9 mg/L, 25.6 mg/L for conventional sampling, respectively. The area under the concentration-time curve from 0 to 6 h (AUC0-6h) ranged between 94.8 and 269 mg/L.h for capillary microsampling and from 106 and 303 mg/L.h for conventional sampling. The comparative study conducted was assessed using three different statistical approaches: Bland-Altman and Passing-Bablok regression analyses and the USFDA criterion for the incurred sample reanalysis. The results of this analysis revealed no significant bias and a strong correlation between both sampling methods, with 95% of the calculated concentrations from the paired plasma samples laying within 20% of difference of the mean. This bridging study verifies that capillary microsampling may serve as an alternative to conventional sampling techniques to support clinical applications for measuring vancomycin concentrations in plasma.

Toxicokinetics of Deltamethrin: Dosage Dependency, Vehicle Effects, and Low-Dose Age-Equivalent Dosimetry in Rats.

There is increasing concern that infants and children may be at increased risk of neurological effects of pyrethroids, the most widely used class of insecticide. The objectives of this investigation were to (1) characterize the dose-dependent toxicokinetics (TK) of deltamethrin (DLM) for exposures ranging from environmentally relevant to acutely toxic; (2) determine the influence of an aqueous versus oil vehicle on oral absorption and bioavailability; and (3) determine whether DLM exhibits low-dose, age-equivalent internal dosimetry. Serial arterial plasma samples were obtained for 72 h from adult, male Sprague Dawley rats given 0.05-5.0 mg DLM/kg as an oral bolus in corn oil (CO). DLM exhibited linear, absorption rate-limited TK. Increases in maximum plasma concentration (Cmax) and AUC∘∞ were directly proportional to the dose. Oral bioavailability was quite limited. The vehicle and its volume had modest effect on the rate and extent of systemic absorption in adult rats. Postnatal day (PND) 15, 21, and 90 (adult) rats received 0.10, 0.25, or 0.50 mg DLM/kg orally in CO and were sacrificed periodically for plasma, brain, and liver collection. Age-dependent differences between PND 15 and 90 plasma Cmax and AUC∘24 values progressively diminished as the dose decreased, but there was a lack of low dose age equivalence in these brain and liver dosimeters. Other maturational factors may account for the lack of the low-dose age equivalence in brain and liver. This investigation provides support for the premise that the relatively low metabolic capacity of immature subjects may be adequate to effectively eliminate trace amounts of DLM and other pyrethroids from the plasma.

Dietary supplementation of branched-chain amino acids increases muscle net amino acid fluxes through elevating their substrate availability and intramuscular catabolism in young pigs.

Branched-chain amino acids (BCAA) have been clearly demonstrated to have anabolic effects on muscle protein synthesis. However, little is known about their roles in the regulation of net AA fluxes across skeletal muscle in vivo. This study was aimed to investigate the effect and related mechanisms of dietary supplementation of BCAA on muscle net amino acid (AA) fluxes using the hindlimb flux model. In all fourteen 4-week-old barrows were fed reduced-protein diets with or without supplemental BCAA for 28 d. Pigs were implanted with carotid arterial, femoral arterial and venous catheters, and fed once hourly with intraarterial infusion of p-amino hippurate. Arterial and venous plasma and muscle samples were obtained for the measurement of AA, branched-chain α-keto acids (BCKA) and 3-methylhistidine (3-MH). Metabolomes of venous plasma were determined by HPLC-quadrupole time-of-flight-MS. BCAA-supplemented group showed elevated muscle net fluxes of total essential AA, non-essential AA and AA. As for individual AA, muscle net fluxes of each BCAA and their metabolites (alanine, glutamate and glutamine), along with those of histidine, methionine and several functional non-essential AA (glycine, proline and serine), were increased by BCAA supplementation. The elevated muscle net AA fluxes were associated with the increase in arterial and intramuscular concentrations of BCAA and venous metabolites including BCKA and free fatty acids, and were also related to the decrease in the intramuscular concentration of 3-MH. Correlation analysis indicated that muscle net AA fluxes are highly and positively correlated with arterial BCAA concentrations and muscle net BCKA production. In conclusion, supplementing BCAA to reduced-protein diet increases the arterial concentrations and intramuscular catabolism of BCAA, both of which would contribute to an increase of muscle net AA fluxes in young pigs.

Transcriptome analysis reveals non-identical microRNA profiles between arterial and venous plasma.

Circulating microRNAs presented in venous plasma have been demonstrated as powerful biomarkers for the complex diseases like cancer. Nevertheless, those presented in arterial plasma remained largely unexplored. Here, using microarray technique, we compared microRNA expression profiles of the matched arterial and venous plasma samples from the same male rats. Though the microRNA profiles were largely similar, we identified 24 differentially expressed microRNAs, including 10 arterial highly expressed microRNAs and 14 venous highly expressed microRNAs. The differentially expressed microRNAs were validated by qRT-PCR. Computational analysis of these microRNAs and their targets indicated that arterial highly expressed microRNAs were overrepresented for functional terms like hematopoiesis and diseases like Crohn's Disease and leukemia; while venous highly expressed microRNAs were enriched for cell differentiation function, and diseases like distal myopathies and heart failure. Our analysis also suggested significant correlations between plasma microRNA expression and tissue microRNA expression. Four arterial highly expressed microRNAs also showed enriched expression in specific tissues and would be novel biomarker candidates.

Living with lipids: Reflections of a fat physiologist

Living with lipids: Reflections of a fat physiologist