Fentanyl dosage for preterm infants suggested by a pharmacokinetic, -dynamic, and -genetic model.

BACKGROUND: Fentanyl is commonly administered for procedural pain management in preterm infants, but target concentrations have not yet been defined. METHODS: To investigate pharmacokinetics (PK), -dynamics (PD), and -genetics (PG), 25 infants (gestational age 23.3-34.1 weeks) received a fentanyl dose before a skin-breaking procedure (0.5 µg/kg) or tracheal intubation (2 µg/kg). Four pain scales were used as a PD endpoint to evaluate efficacy. The impact of polymorphism in genes encoding enzymes (UGT2B7, CYP3A7, CYP3A4, COMT, CYP2D6, KCNJ6), transporters (SLC22A1, ABCC1, ABCC3) and receptor (OPRM1) on PK parameters was explored. RESULTS: A two-compartment PK model adequately described the fentanyl concentration. The effects of weight and maturity on the clearance were included as covariates in the model. One genetic variant encoding the ABCC1 transporter (rs111517339 T/TA) and two encoding the ABCC3 transporter (rs11079921 T/C and rs8077268 C/T) had a significant effect on fentanyl elimination that explained 15% of the interindividual variability on the clearance. A proportional odds PK/PD model was used to describe the concentration-effect relationship of fentanyl using the Échelle de douleur et d'inconfort du nouveau-né (EDIN) pain score. CONCLUSION: The simulations suggest that an intravenous dose of 2 µg/kg would be appropriate in preterm infants for a clearly painful procedure, such as an intubation. IMPACT: Design of personalized analgesia with fentanyl for newborn infants should consider maturation and genetic variants of opioid transporters affecting drug elimination. The results indicate that an intravenous dose of 2 µg/kg fentanyl would be suitable before a clearly painful procedure in preterm infants. Genetic variants encoding ABCC1 and ABCC3 transporters increase the clearance of fentanyl, which is a novel finding.

Depression of cortical neuronal activity after a low-dose fentanyl in preterm infants.

AIM: Opioids might be harmful to the developing brain and dosing accuracy is important. We aimed at investigating fentanyl effects on cortical activity in infants using computational re-analysis of bedside recorded EEG signals. METHODS: Fifteen infants born at median 26.4 gestational weeks (range 23.3-34.1), with a birth weight 740 grams (530-1420) and postnatal age 7 days (5-11) received fentanyl 0.5 or 2 µg/kg intravenously before a skin-breaking procedure or tracheal intubation, respectively. Cortical activity was continuously recorded using amplitude-integrated electroencephalography (aEEG).  Analyses using three computational EEG features representing cortical synchrony and signal power, were conducted five minutes pre- and 10 minutes post the drug administration. RESULTS: Visual assessment of trends displayed from the EEG metrics did not indicate systematic changes. However, the magnitude of the changes in the parietal and right hemisphere signals after the dose was significantly correlated (ρ < -0.5, p < 0.05) to the EEG amplitude and frequency power level before drug administration. This effect started after 3-4 min. CONCLUSION: Fentanyl, even in small doses, may affect cortical activity in the preterm brain. The effect is robustly related to the state of cortical activity prior to drug treatment, which must be taken into account when analysing the effects of sedative drugs.

Perinatal risk factors for developmental coordination disorder in children born extremely preterm.

AIM: Children born extremely preterm frequently have developmental coordination disorder (DCD). We aimed to evaluate perinatal risk factors for DCD. METHODS: Swedish national cohort study including 226 children born before 27 gestational weeks without major neurodevelopmental disabilities at 6.5 years. Outcome was DCD, defined as ≤5th percentile on the Movement Assessment Battery for Children-Second Edition. Perinatal risk factors were evaluated using multivariable logistic regression. RESULTS: DCD was present in 84/226 (37.2%) children. Of the risk factors known at 40 weeks gestation, independent and significant risk factors for DCD were: mother's age at delivery (odds ratio [OR] 1.73, 95% confidence interval [CI] 1.07-2.80); pre-eclampsia (2.79, 1.14-6.80); mother born in a non-Nordic country (2.23, 1.00-4.99); gestational age per week increase (0.70, 0.50-0.99) and retinopathy of prematurity (2.48, 1.26-4.87). Of factors known at discharge, postnatal steroids exposure (2.24, 1.13-4.46) and mechanical ventilation (1.76, 1.06-2.09) were independent risk factors when added to the model in separate analyses. CONCLUSION: The risk of DCD in children born extremely preterm was multifactorial and associated with gestational age largely mediated by ROP, maternal factors, pre-eclampsia, administration of postnatal steroids and mechanical ventilation. These risk factors are common among children born extremely preterm, contributing to their high risk of DCD.

No impact of parental singing during the neonatal period on cognition in preterm-born children at 2-3 years.

AIM: Studies examining the long-term effects of neonatal music interventions on the cognition of children born preterm are scarce. We investigated whether a parental singing intervention before term age improves cognitive and language skills in preterm-born children. METHODS: In this longitudinal, two-country Singing Kangaroo, randomised controlled trial, 74 preterm infants were allocated to a singing intervention or control group. A certified music therapist supported parents of 48 infants in the intervention group to sing or hum during daily skin-to-skin care (Kangaroo care) from neonatal care until term age. Parents of 26 infants in the control group conducted standard Kangaroo care. At 2-3 years of corrected age, the cognitive and language skills were assessed with the Bayley Scales of Infant and Toddler Development, Third Edition. RESULTS: There were no significant differences in cognitive and language skills between the intervention and control groups at the follow-up. No associations between the amount of singing and the cognitive and language scores were found. CONCLUSION: Parental singing intervention during the neonatal period, previously shown to have some beneficial short-term effects on auditory cortical response in preterm infants at term age, showed no significant long-term effects on cognition or language at 2-3 years of corrected age.

A sensitive assay for dNTPs based on long synthetic oligonucleotides, EvaGreen dye and inhibitor-resistant high-fidelity DNA polymerase.

Deoxyribonucleoside triphosphates (dNTPs) are vital for the biosynthesis and repair of DNA. Their cellular concentration peaks during the S phase of the cell cycle. In non-proliferating cells, dNTP concentrations are low, making their reliable quantification from tissue samples of heterogeneous cellular composition challenging. Partly because of this, the current knowledge related to the regulation of and disturbances in cellular dNTP concentrations derive mostly from cell culture experiments with little corroboration at the tissue or organismal level. Here, we fill the methodological gap by presenting a simple non-radioactive microplate assay for the quantification of dNTPs with a minimum requirement of 4-12 mg of biopsy material. In contrast to published assays, this assay is based on long synthetic single-stranded DNA templates (50-200 nucleotides), an inhibitor-resistant high-fidelity DNA polymerase, and the double-stranded-DNA-binding EvaGreen dye. The assay quantified reliably less than 50 fmol of each of the four dNTPs and discriminated well against ribonucleotides. Additionally, thermostable RNAse HII-mediated nicking of the reaction products and a subsequent shift in their melting temperature allowed near-complete elimination of the interfering ribonucleotide signal, if present. Importantly, the assay allowed measurement of minute dNTP concentrations in mouse liver, heart and skeletal muscle.

Changes in pulmonary oxygen content are detectable with laser absorption spectroscopy: proof of concept in newborn piglets.

BACKGROUND: Using an optical method based on tunable diode laser absorption spectroscopy, we previously assessed oxygen (O2) and water vapor (H2O) content in a tissue phantom of the preterm infant lung. Here we applied this method on newborn piglets with induced lung complications. METHODS: Five mechanically ventilated piglets were subjected to stepwise increased and decreased fraction of inspired oxygen (FiO2), to atelectasis using a balloon catheter in the right bronchus, and to pneumothorax by injecting air in the pleural cavity. Two diode lasers (764 nm for O2 gas absorption and 820 nm for H2O absorption) were combined in a probe delivering light either externally, on the skin, or internally, through the esophagus. The detector probe was placed dermally. RESULTS: Calculated O2 concentrations increased from 20% (IQR 17-23%) when ventilated with room air to 97% (88-108%) at FiO2 1.0. H2O was only detectable with the internal light source. Specific light absorption and transmission patterns were identified in response to atelectasis and pneumothorax, respectively. CONCLUSIONS: The optical method detected FiO2 variations and discriminated the two induced lung pathologies, providing a rationale for further development into a minimally invasive device for real-time monitoring gas changes in the lungs of sick newborn infants. IMPACT: Optical spectroscopy can detect pulmonary complications in an animal model. Oxygen concentrations can be evaluated in the lungs. Presents a novel minimally invasive method to detect lung oxygenation and complications. Potential to be developed into a lung monitoring method in newborn infants. Potential for bed-side detection of pulmonary complications in newborn infants.

Difficult birth is the main contributor to birth-related fracture and accidents to other neonatal fractures.

AIM: Specific birth-related fractures have been studied; underestimates might be a problem. We aimed to assess all fractures diagnosed as birth-related as well as other neonatal fractures. METHODS: A population-based study on all infants born in Sweden 1997-2014; data were retrieved from the Swedish Health Registers (10th version of International Classification of Diseases. Outcome measures were birth-related fractures (ICD-10 P-codes) and other neonatal fractures (ICD-10 S-codes). RESULTS: The overall fracture incidence was 2.9 per 1000 live birth (N = 5336); 92.6% had P-codes and 7.4% (S-codes). Some birth-related fractures were diagnosed beyond the neonatal period. Other neonatal fractures could have been birth-related. Clavicle fracture (88.8%) was associated with adverse maternal and infant anthropometrics and birth complications. The few neonates with rib fractures all had concomitant clavicle fracture. For skull fractures, a minor part was birth-related and most were associated with accidents. Half of the long bone fractures were associated with accidents. Birth-related femur fractures were associated with bone fragility risk factors. Five infants with abuse diagnoses had fractures: skull (4), long bone (2) and rib (1). CONCLUSION: Birth-related and other neonatal fractures are rarely diagnosed. Difficult birth is the main contributor to birth-related fracture and accidents to other neonatal fractures.

NAD+ repletion produces no therapeutic effect in mice with respiratory chain complex III deficiency and chronic energy deprivation.

Biosynthetic precursors of NAD+ can replenish a decreased cellular NAD+ pool and, supposedly via sirtuin (SIRT) deacetylases, improve mitochondrial function. We found decreased hepatic NAD+ concentration and downregulated biosynthesis in Bcs1lp.S78G knock-in mice with respiratory chain complex III deficiency and mitochondrial hepatopathy. Aiming at ameliorating disease progression via NAD+ repletion and improved mitochondrial function, we fed these mice nicotinamide riboside (NR), a NAD+ precursor. A targeted metabolomics verified successful administration and suggested enhanced NAD+ biosynthesis in the treated mice, although hepatic NAD+ concentration was unchanged at the end point. In contrast to our expectations, NR did not improve the hepatopathy, hepatic mitochondrial respiration, or survival of Bcs1lp.S78G mice. We linked this lack of therapeutic effect to NAD+-independent activation of SIRT-1 and -3 via AMPK and cAMP signaling related to the starvation-like metabolic state of Bcs1lp.S78G mice. In summary, we describe an unusual metabolic state with NAD+ depletion accompanied by energy deprivation signals, uncompromised SIRT function, and upregulated oxidative metabolism. Our study highlights that the knowledge of the underlying complex metabolic alterations is critical when designing therapies for mitochondrial dysfunction.-Purhonen, J., Rajendran, J., Tegelberg, S., Smolander, O.-P., Pirinen, E., Kallijärvi, J., Fellman, V. NAD+ repletion produces no therapeutic effect in mice with respiratory chain complex III deficiency and chronic energy deprivation.

Population-based register study of children born in Sweden from 1997 to 2014 showed an increase in rickets during infancy.

AIM: This population-based study assessed the incidence of rickets in infants up to age of one born in Sweden from 1997 to 2014. We also examined maternal and perinatal factors and co-morbidity. METHODS: We used Swedish National Board of Health and Welfare registers and data from Statistics Sweden. The outcome measure was an International Classification of Diseases, Tenth Revision, code for rickets. RESULTS: There were 273 cases of rickets, with an incidence of 14.7 per 100 000 and a 10-fold incidence increase between 1997 and 2014. The majority (78.4%) were born preterm, half were small-for-gestational age (SGA) (birthweight <10th percentile), 4.8% were born to Asian-born mothers and 3.5% to African-born mothers. The adjusted odds ratios by birth week were 182 (95% CI: 121-272) before 32 weeks and 10.8 (95% CI: 6.72-17.4) by 32-36 weeks. Preterm infants with necrotising enterocolitis had very high odds for rickets and so did SGA term-born infants and those born to African-born mothers. The odds for rickets among preterm infants increased considerably during the later years. CONCLUSION: Rickets increased 10-fold in Sweden from 1997 to 2014 and was mainly associated with prematurity, SGA and foreign-born mothers. Possible reasons may include increased preterm survival rates and improved clinical detection and registration.

Individual variations in fentanyl pharmacokinetics and pharmacodynamics in preterm infants.

AIM: Fentanyl pharmacokinetics and pharmacodynamics are lacking in preterm infants. Our aim was to study these and their relation with a new formulation of fentanyl 5 µg/mL for procedural pain. METHODS: Preterm infants were given 0.5 (n = 20, median gestational age 26.5; range 23.3-34.1 weeks) and 2 µg/kg (n = 8, 27.4; 25.3-30.7 weeks) fentanyl, respectively, before skin-breaking procedures or tracheal intubation. Blood samples were collected after ten minutes, two, four, eight and 24 hours. Physiologic parameters were monitored and pain scores assessed. RESULTS: The median fentanyl concentrations were 0.18, 0.15, 0.15 and 0.57, 0.37, 0.35 ng/mL at 15-31 minutes, two and four hours and the half-lives were 1.6 to 20.5 or 4.1 to 32.6 hours for the low- and high-dose groups, respectively. A significant correlation was seen between weight at study inclusion and half-life (Spearman's r = -0.9, p < 0.001), volume of distribution (r = -0.8, p < 0.01) and clearance (r = -0.9, p < 0.01) in the low-dose group (n = 9). Pain assessment results were not correlated to pharmacokinetic variables. Fentanyl was well tolerated. CONCLUSION: The inter-individual variation of fentanyl pharmacokinetics is large in preterm infants, and the dose of 0.5 µg/kg seems not effective for skin-breaking procedures.

Oxygen consumption in platelets as an adjunct diagnostic method for pediatric mitochondrial disease.

BackgroundDiagnosing mitochondrial disease (MD) is a challenge. In addition to genetic analyses, clinical practice is to perform invasive procedures such as muscle biopsy for biochemical and histochemical analyses. Blood cell respirometry is rapid and noninvasive. Our aim was to explore its possible role in diagnosing MD.MethodsBlood samples were collected from 113 pediatric patients, for whom MD was a differential diagnosis. A respiratory analysis model based on ratios (independent of mitochondrial specific content) was derived from a group of healthy controls and tested on the patients. The diagnostic accuracy of platelet respirometry was evaluated against routine diagnostic investigation.ResultsMD prevalence in the cohort was 16%. A ratio based on the respiratory response to adenosine diphosphate in the presence of complex I substrates had 96% specificity for disease and a positive likelihood ratio of 5.3. None of the individual ratios had sensitivity above 50%, but a combined model had 72% sensitivity.ConclusionNormal findings of platelet respirometry are not able to rule out MD, but pathological results make the diagnosis more likely and could strengthen the clinical decision to perform further invasive analyses. Our results encourage further study into the role of blood respirometry as an adjunct diagnostic tool for MD.

Preterm arteries in childhood: dimensions, intima-media thickness, and elasticity of the aorta, coronaries, and carotids in 6-y-old children born extremely preterm.

BACKGROUND: Preterm birth increases risk for adult cardiovascular disease. We hypothesized that arteries in 6-y-old children born preterm are narrower, with thicker intima-media and stiffer than in peers born at term. METHODS: Children born extremely preterm (EXP, n = 176, birthweights: 348-1,161 g) and at term (CTRL, n = 174, birthweights: 2,430-4,315 g) were included. Using ultrasonography, we determined diameters of the coronaries (CA), common carotid arteries (CCA) and aorta, the carotid intima media thickness (cIMT), and the stiffness index of the CCA and aorta. RESULTS: Arteries were 5-10% narrower in EXP than in CTRL (P < 0.005) but after adjustment for body surface area, diameter differences diminished or disappeared. EXP-children born small for gestational age exhibited similar arterial dimensions as those born appropriate for date. The cIMT was 0.38 (SD = 0.04) mm and did not differ between groups. Carotid but not aortic stiffness was lower in EXP than in CTRL. CONCLUSION: In 6-y-old children born extremely preterm, conduit arteries are of similar or smaller size than in controls born at term, and they have no signs of accelerated intima media thickening or arterial stiffening. While these findings are reassuring for these children and their families, the causal pathways from preterm birth to adult cardiovascular disease remain unknown.

Diode laser spectroscopy for noninvasive monitoring of oxygen in the lungs of newborn infants.

BACKGROUND: Newborn infants may have pulmonary disorders with abnormal gas distribution, e.g., respiratory distress syndrome. Pulmonary radiography is the clinical routine for diagnosis. Our aim was to investigate a novel noninvasive optical technique for rapid nonradiographic bedside detection of oxygen gas in the lungs of full-term newborn infants. METHODS: Laser spectroscopy was used to measure contents of oxygen gas (at 760 nm) and of water vapor (at 937 nm) in the lungs of 29 healthy newborn full-term infants (birth weight 2,900-3,900 g). The skin above the lungs was illuminated using two low-power diode lasers and diffusely emerging light was detected with a photodiode. RESULTS: Of the total 390 lung measurements performed, clear detection of oxygen gas was recorded in 60%, defined by a signal-to-noise ratio of >3. In all the 29 infants, oxygen was detected. Probe and detector positions for optimal pulmonary gas detection were determined. There were no differences in signal quality with respect to gender, body side or body weight. CONCLUSION: The ability to measure pulmonary oxygen content in healthy full-term neonates with this technique suggests that with further development, the method might be implemented in clinical practice for lung monitoring in neonatal intensive care.

Genetic Predisposition to Poor Opioid Response in Preterm Infants: Impact of KCNJ6 and COMT Polymorphisms on Pain Relief After Endotracheal Intubation.

BACKGROUND: Single-nucleotide polymorphisms in genes involved in pain control might predispose to exaggerated sensitivity or difference in opioid analgesic effect. The relevance of the KCNJ6 -1250G>A (rs6517442, c.-1787G>A) and the catecholamine-O-methyltransferase (COMT) c.472G>A (rs4680, ValMet) single-nucleotide polymorphisms were studied in preterm infants needing intubation and randomized to a premedication strategy including remifentanil (n = 17) or morphine (n = 17). METHODS: Pain was scored with Astrid Lindgren and Lund Children's Hospital Pain Assessment Scale every 30 minutes for 6 hours. The pain relief provided by the opioids was compared between the different KCNJ6 and COMT genotypes. RESULTS: Infants homozygous for the KCNJ6 -1250A allele had an increased duration after intubation to achieve a score indicating no pain compared with infants with the A/G or G/G genotypes (182 ± 30, 109 ± 29, and 60 ± 21 minutes, respectively; Logrank = 7.5, P = 0.006). Similarly, the duration was increased in individuals with the COMT Val/Val alleles compared with Val/Met and Met/Met (285 ± 37, 137 ± 25, and 63 ± 15 minutes, respectively; Logrank = 14.4, P = 0.0021). Cox proportional hazards analysis confirmed that the variation in both genes was independently associated with susceptibility to respond to therapy. CONCLUSION: We conclude that the KCNJ6 -1250A and COMT Val alleles are predisposing preterm newborns to diminished opioid-induced pain relief.

Effect of High-Carbohydrate Diet on Plasma Metabolome in Mice with Mitochondrial Respiratory Chain Complex III Deficiency.

Mitochondrial disorders cause energy failure and metabolic derangements. Metabolome profiling in patients and animal models may identify affected metabolic pathways and reveal new biomarkers of disease progression. Using liver metabolomics we have shown a starvation-like condition in a knock-in (Bcs1lc.232A>G) mouse model of GRACILE syndrome, a neonatal lethal respiratory chain complex III dysfunction with hepatopathy. Here, we hypothesized that a high-carbohydrate diet (HCD, 60% dextrose) will alleviate the hypoglycemia and promote survival of the sick mice. However, when fed HCD the homozygotes had shorter survival (mean ± SD, 29 ± 2.5 days, n = 21) than those on standard diet (33 ± 3.8 days, n = 30), and no improvement in hypoglycemia or liver glycogen depletion. We investigated the plasma metabolome of the HCD- and control diet-fed mice and found that several amino acids and urea cycle intermediates were increased, and arginine, carnitines, succinate, and purine catabolites decreased in the homozygotes. Despite reduced survival the increase in aromatic amino acids, an indicator of liver mitochondrial dysfunction, was normalized on HCD. Quantitative enrichment analysis revealed that glycine, serine and threonine metabolism, phenylalanine and tyrosine metabolism, and urea cycle were also partly normalized on HCD. This dietary intervention revealed an unexpected adverse effect of high-glucose diet in complex III deficiency, and suggests that plasma metabolomics is a valuable tool in evaluation of therapies in mitochondrial disorders.

Auditory event-related potentials are related to cognition at preschool age after very preterm birth.

BACKGROUND: Auditory event-related potentials (AERP) are neurophysiological correlates of sound perception and cognitive processes. Our aim was to study in very preterm born children at preschool age if AERP correlate with cognitive outcome. METHODS: Seventy children (mean ± SD gestational age 27.4 ± 1.9 wk, birth weight 996 ± 288 g) were investigated at age 4.3-5.3 y with psychological testing (WPPSI-R, four subtests of NEPSY). Electroencephalogram was recorded while they listened to a repeated standard tone, randomly replaced by one of three deviants. Latencies and amplitudes for AERP components and mean amplitudes in successive 50-ms AERP time windows were measured. RESULTS: Better cognitive test results and higher gestational age correlated with shorter P1 latencies and more positive mean amplitudes 150-500 ms after stimulus change onset. Neonatal brain damage was associated with a negative displacement of AERP curves. Neonatal morbidity had an impact on earlier time windows while gestational age and brain damage on both early and later time windows. CONCLUSION: AERP measures were associated with cognitive outcome. Neonatal morbidity mainly affects early cortical auditory encoding, while immaturity and brain damage additionally influence higher cortical functions of auditory perception and distraction. Perinatal auditory environment might play a role in development of auditory processing.

Extremely preterm birth affects boys more and socio-economic and neonatal variables pose sex-specific risks.

AIM: The early identification of at-risk extremely preterm (EPT) children could improve long-term outcomes. This study sought to investigate sex differences in developmental outcomes and to identify sex-specific predictors at two and a half years of age. METHODS: We assessed 217 boys and 181 girls born before 27-week gestation using the Bayley Scales of Infant and Toddler Development, third edition (Bayley-III), as a part of the Extremely Preterm Infants in Sweden Study. Sex-specific differences were calculated. Socio-economic, birth and neonatal factors were calculated separately for boys and girls using regression models. RESULTS: Girls scored significantly higher than boys on all Bayley-III indices. In both sexes, brain injury, long-term ventilator treatment and foreign-born mothers predicted lower scores. Receiving breast milk by hospital discharge predicted higher scores. Severe retinopathy of prematurity was the strongest predictor of cognitive and language deficits in boys. High parental education predicted higher cognitive and language scores in girls, whereas severe bronchopulmonary dysplasia was the strongest predictor of motor deficits. CONCLUSION: Extreme prematurity seems to affect boys more than girls. Socio-economic and neonatal factors confer similar risks or protections on both sexes, but some variables pose sex-specific risks. An awareness of risk factors may provide the basis for treatment and follow-up guidelines.

Parents tend to underestimate cognitive deficits in 10- to 13-year-olds born with an extremely low birth weight.

AIM: This study determined the cognitive outcomes of Finnish children born with an extremely low birth weight (ELBW) and assessed the agreement between their neuropsychological assessment and how their parents evaluated their cognitive difficulties. METHODS: The study focused on 121 children from an ELBW cohort with a mean age of 11.6 years (range 10.3-13.8) and assessed them using a standardised test of intelligence, a neuropsychological test battery and a parental developmental questionnaire. The results were compared with the test norms. RESULTS: ELBW children exhibited global cognitive impairment compared to the test norms, with no differences between children who were small or appropriate for gestational age. Children with average intelligence displayed specific impairment in executive, sensorimotor and visuospatial functions. Corresponding functions in the parental evaluation and neuropsychological assessment were associated, but 16-26% of children scoring under the clinical cut-off value in the neuropsychological test domains were not detected by the parental evaluations. CONCLUSION: Children born with an ELBW faced a high risk of global cognitive impairment at a mean age of 11.6 years, and those with average intelligence were at risk of specific cognitive sequelae. Compared to the neuropsychological tests, up to one-fourth of the parents underestimated their child's cognitive problems.