Molecular profiling of preterm infants' urine offers a promising noninvasive strategy to identify biomarkers. In this prospective study, urine samples were collected from 64 preterm infants ( < 32 weeks gestation) on postnatal days 1-4, 6, 8, 28 and at term-equivalent age. Forty-three metabolites and neurotransmitters were quantified by LC-MS/MS. Brain injury severity was quantified using the MRI-derived Global Brain Abnormality Score, capturing white matter, cortex, deep grey matter, and cerebellar injury. Distinct temporal patterns of urinary metabolites were associated with global brain injury severity. Early energy-related elevations (lactate, α-hydroxybutyrate) were followed by broader alterations in amino acid, fatty acid, and tryptophan-kynurenine pathways between postnatal days 3-6, defining a promising neurosurveillance window that may implicate mitochondrial dysfunction, oxidative stress, and excitotoxicity as relevant injury mechanisms. At the default decision threshold, multi-block modeling identified 100% of severe injury patients (AUC = 0.849) using day 3-6 data. Exploratory prediction of region-specific MRI anomaly was most robust for cerebellum (AUC = 0.891; full model) and white matter (AUC = 0.724; early model). Urine metabolomics may detect evolving preterm brain injury before MRI at term-equivalent age, with notable results between days 3 and 6. Combining metabolomic insights with clinical and imaging data could allow earlier risk assessment and neuroprotective treatment in preterm infants. IMPACT: Serial urine metabolomics in very preterm infants captures dynamic metabolic disturbances associated with brain injury. Urine patterns show possibly region-specific brain vulnerability. Days 3-6 define a critical window for stratifying risk of severe brain injury at term-equivalent age, shifting focus toward earlier care decisions.

OBJECTIVES:Urinary creatinine concentration is widely used to normalize urinary analytes. However, its reliability in preterm neonates remains uncertain. This study aimed to evaluate urinary creatinine concentrations in very low gestational age (VLGA) neonates and assess its impact on the interpretation of urinary creatinine normalized urinary amino acids. METHODS:Spot urine samples were collected from VLGA neonates (22-31 weeks gestational age, n = 32) and full-term neonates (≥ 37 weeks, n = 22) during the first days of life. Urinary creatinine was measured serially, and biological variation (within- and between-subject) was assessed. Urinary amino acids were measured on the 4th day in full-term neonates and in VLGA neonates at term-equivalent age using LC-MS/MS and expressed both as absolute concentrations (µmol/l) and normalized to creatinine (mmol/mol creatinine). RESULTS:Urinary creatinine concentrations were significantly lower in VLGA neonates compared to full-term neonates during the first days of life (p ≤ 0.004). A positive correlation between birth weight and urinary creatinine was observed (r = 0.46, p = 0.002). Within-subject biological variation was lower in VLGA neonates, while between-subject variation was comparable between groups. Urinary amino acid concentrations expressed in μmol/l did not differ significantly between groups, however, when expressed relative to creatinine, multiple significant differences emerged. CONCLUSIONS:Gestational age should be considered when interpreting creatinine normalized urinary amino acids concentrations in neonates.

D-dimer as a mixture of soluble cross-linked fibrin degradation products cleaved by plasmin isa well-established biomarker of in vivo blood coagulation activation with concomitant secondaryfibrinolysis that is widely determined in a broad variety of clinical settings. D-dimer interpretationis hampered by the lack of an assay international standard and assay harmonization across laboratories,preanalytical variables affecting the results, multiple assays available with different units,and the impact of heterophilic nonspecific IgG or IgM antibodies, which cause falsely increased ordecreased results. Different reactivity of monoclonal antibodies against D-dimer molecule results in variability of results obtained with different assays within individual patients. Compelling clinical evidence supports a high negative predictive value of age-adjusted D-dimer concentration in rulingout venous thromboembolism - both pulmonary embolism and deep-vein thrombosis. ElevatedD-dimer levels are observed in acute myocardial infarction or stroke, aortic aneurysm and acute dissection,atrial fibrillation, heart failure, following invasive procedures, and during infection. Elevated D-dimer can be found in chronic venous insufficiency, chronic inflammatory diseases, cancer, sepsis,autoimmune disorders, diabetes, advanced age, strenuous physical activity, and pregnancy. Althoughelevated D-dimer is associated with increased risk of first and recurrent venous thromboembolism,it is not per se an indication for anticoagulant treatment, and it should lead to in-depth diagnosticevaluation to identify mechanisms underlying such abnormality. The current overview summarizesboth laboratory and clinical knowledge on D-dimer measurement and interpretation with a focuson acute and chronic cardiovascular diseases.

Obesity, regardless of age, is associated with various metabolic disorders. Amino acids, as essential food ingredients, play a critical role in regulating glucose and energy metabolism. Thus, their plasma concentrations may reflect the metabolic state of the individual. This narrative review aims to find out similarities in plasma amino acid profiles across the selected obese individuals (children, adolescents, and adults). An analysis of 21 original studies revealed similar amino acid alterations regardless of the age group— in particular, elevated levels of branched-chain amino acids (BCAAs: isoleucine, leucine, valine), tyrosine, glutamic acid, alanine, and proline, as well as decreased concentrations of glutamine, serine, glycine, and asparagine. These findings suggest the presence of a common, age-independent metabolic signature of amino acids in obesity. Such changes may reflect impaired amino acid catabolism and have been linked to insulin resistance, increased risk of type 2 diabetes risk, and other metabolic complications. There is a need to establish age- and population-specific reference values for plasma amino acids to use amino acid profiling as a diagnostic and prognostic tool management of obesity. Accurate plasma amino acid profile measurements could support early detection of obesity-related metabolic disturbances and the implementation of targeted nutritional and therapeutic strategies.

Introduction:This study aimed to assess the associationbetween the urinary lactate-to-creatinine ratio (ULCR) andbrain spectroscopy (1H-MRS)findings in very low gestationalage (VLGA) infants with and without preterm brain injury.Methods:Urine samples were collected from 54 VLGA in-fants during thefirst week of life, after 1 month of life, and atterm-equivalent age (TEA). Urinary lactate was measured viahighly selective liquid chromatography-tandem massspectrometry (LC-MS/MS) with a quantitative organic acidanalysis kit and expressed as the ULCR. Magnetic resonanceimaging and1H-MRS were performed at TEA. The Kidokorograding system was used to assess the Global Brain Ab-normality Score (GBAS).Results:VLGA infants with a GBASmoderate + severe had higher ULCRs on the 2nd and 3rddays of life (DOLs) than those with a GBAS normal or mild.Only the GBAS moderate + severe subgroup presented witha secondary increase in the ULCR on the 3rd DOL, whereas inthe GBAS normal or mild, the ULCR oscillated around similarvalues or gradually decreased. Significant positive correla-tions were detected between the ULCR on the 3rd DOL andthe lactate/creatinine and lactate/N-acetyl aspartate ratiosmeasured via1H-MRS at TEA (r= 0.308;p= 0.022 andr=0.334;p= 0.013, respectively).Conclusions:An increasedULCR during thefirst 3 DOLs in patients with a GBASmoderate + severe suggest an energy catastrophe that mayplay a role in the development of premature brain injury.Serial measurement of the ULCR during thefirst DOLs mayhelp in the early identification of premature infants at risk formoderate + severe brain damage.(c) 2024 The Author(s).Published by S. Karger AG, Basel.

Antibiotic therapy requires appropriate dosage of drugs for effective treatment. Too low antibiotic concentrations may lead to treatment failure and the development of resistant pathogens, whereas overdosing may cause neurological side effects or hemolytic diseases. Meropenem and linezolid are used only in the treatment of serious infections or when other antibiotics are no longer effective as well as for treating central nervous system infections. It is difficult or sometimes even impossible to predict the relation between dosing of antibiotics and its cerebrospinal fluid (CSF) concentration; thus, a method of determining antibiotics not only in the blood but also in the CSF is needed. Analytical method validation is an integral part of good laboratory practice and ensures high accuracy of the results. We performed complete validation process according to the Food and Drug Administration and European Medicine Agency, covering the aspects precision, specificity, accuracy, recovery, limit of detection, limit of quantification, stability, carry-over, and matrix effects. Our liquid chromatography-tandem mass spectrometry method for the simultaneous measurement of meropenem and linezolid in different matrix meets all the acceptance criteria. The method was successfully applied to determine meropenem and linezolid concentrations in serum and CSF samples obtained from children treated with these antibiotics.

The role of amino acids in cholesterol gallstone formation is not known. Therefore, the aim of the study was to determine the amino acid profile in the bile of patients with and without cholecystolithiasis in relation to bile lithogenicity and telocyte numbers within the gallbladder wall. The study included 23 patients with cholecystolithiasis and 12 gallstone-free controls. The levels of free amino acids in the bile were measured, and telocytes were identified and quantified in the gallbladder muscle wall. The mean values of valine, isoleucine, threonine, methionine, phenylalanine, tyrosine, glutamic acid, serine alanine, proline and cystine were significantly higher in the study group than in the controls (p from 0.0456 to 0.000005), and the mean value of cystine was significantly lower in patients with gallstone disease than in the controls (p = 0.0033). The relationship between some of the amino acids, namely alanine, glutamic acid, proline, cholesterol saturation index (CSI) and the number of telocytes was significant (r = 0.5374, p = 0.0051; r = 0.5519, p = 0.0036; and r = 0.5231, p = 0.0071, respectively). The present study indicates a potential relationship between the altered amino acid composition of bile and the reduced number of telocytes in the gallbladder muscle wall in cholelithiasis.

BackgroundIt is accepted that plasma branched-chain amino acids (BCAAs) and aromatic amino acids (AAAs) are closely related to metabolic risk. Arterial hypertension, metabolic syndrome, endothelial dysfunction, inflammation, and metabolic dysfunction-associated fatty liver disease (MAFLD) are frequently seen in obese patients. Many attempts have been made to find biochemical indicators for the early detection of metabolic complications in children. It is not known if different amino acid profiles and BCAA and AA concentrations in overweight and obese children correlate with chemerin, proinflammatory, and simple biochemical markers. Thus, the study aimed to find out the early markers of cardiovascular disease and MAFLD in overweight and obese children.Materials and methodsThe study included 20 overweight and obese children (M/F 12/8; mean age 7.7 ± 2.3 years; BMI 26.8 ± 5.0 kg/m2) and 12 non-obese children (control group) (M/F 4/8; mean age 6.5 ± 2.2 years; BMI 14.8 ± 1.5 kg/m2). The following plasma amino acids were measured: aspartic acid, glutamic acid, serine, asparagine, glycine, glutamine, taurine, histidine, citrulline, threonine, alanine, arginine, proline, tyrosine, methionine, valine, isoleucine, leucine, phenylalanine, tryptophan, ornithine, and lysine. Chemerin, high-sensitivity C-reactive protein (hs-CRP), interleukin-6 (IL-6), and basic biochemistry parameters were measured.ResultsThe mean plasma levels of leucine, isoleucine, valine, phenylalanine, tyrosine, glutamic acid, and alanine were significantly higher in overweight and obese children than in the control group (p<0.03–p<0.0004). Conversely, the mean values of serine, asparagine, glutamine, and citrulline were significantly lower in overweight and obese children than in the control group (p<0.03–p<0.0007). Isoleucine, leucine, valine (BCAAs) tyrosine, and phenylalanine (AAAs) levels showed a positive correlation with uric acid, ALT, hs-CRP, and chemerin (r=0.80–0.36; p<0.05-p<0.00001), but not with IL-6. The mean values of glucose, IL-6, hs-CRP, chemerin, uric acid, and ALT were significantly higher in overweight and obese children than in the control group (p<0.03–p<0.00002). In contrast, the lipid profile did not differ between groups.ConclusionAn abnormal amino acid profile in overweight and obese pre-pubertal children, accompanied by elevated ALT and UA observed in the studied cohort, may suggest early metabolic disturbances that can potentially lead to metabolic syndrome, or MAFLD, and increased cardiovascular risk.

Searchable abstracts of presentations at key conferences in endocrinology ISSN 1470-3947 (print) | ISSN 1479-6848 (online)

Background: Serum osmolality can be measured (Omeas) or calculated (Ocal). Many formulas for Ocal have been already published, but data regarding the most accurate equation in small babies is not available. Thus, we aim to compare Omeas and Ocal obtained by different formulas in newborns and small children. Methods: The study included 280 serum samples taken from children, from the first day of life to 2 years (mean age 8.2 ± 7.6 months) treated in the University Children's Hospital in Krakow. The serum osmolality was measured by osmometer and calculated by 8 common formulas. Results: The mean value of Omeas (2 8 5 .8 ± 5 .1 mOsm/kgH2O) was significantly different as compared to the mean values of Ocal (p< 0.01) for all formulas, except Ocal obtained by the formula: 1.86*(N a + K) +1.15*Glu + Urea + 14. According to Bland-Altman analysis, this formula showed the best performance for estimating osmolality. In children under 3 months of life Passing-Bablok regression indicated both systematic and proportional error for results obtained by each formula compared to the measured values. Conclusions: To calculate osmolarity in children aged between 3 months and 2 years old the following equation: 1 .86*(N a + K) + 1.1 5*G lu+ U rea+ 14 might be used, whereas serum osmolality in children up to 3 month of life should be measured.

Introduction The aim of the study was to address the fatty acid (FA) status and its relationship with disease activity in patients with inflammatory bowel disease (IBD). Methods FA levels of the phospholipid fraction in serum and a colon biopsy specimen were measured in 17 patients with IBD. Results A negative correlation between the histological activity of inflammation of the disease and the ratio of polyunsaturated FAs/no polyunsaturated FAs was observed. Moreover, the level of that ratio was lower in patients with IBDs as compared to controls. Conclusions The FA profile in serum and in a colon biopsy specimen in patients with IBD is characteristic for essential fatty acid insufficiency.

INTRODUCTION:The influence of growth hormone (GH) treatment on amino acids (AAs) profile in patients with Turner syndrome (TS) was investigated.MATERIAL AND METHODS:The study group included girls with TS: treated with GH (GH+) and girls with no GH treatment (GH-). The control group consisted of healthy girls. Free plasma AAs were measured by the LC/MS/MS.RESULTS:The plasma concentrations of glutamine, threonine were significantly higher in group GH+ than in group GH- (p < 0.05). In group GH- the values of glutamine, alanine, isoleucine, glutamic acid were significantly different than in the control (p < 0.05-p < 0.008).CONCLUSION:AAs profile in girls with TS might be characteristic for the disease but also depends on GH treatment.

Monitoring of bilirubin concentration is essential during early neonatal life. According to the American Academy of Pediatrics Clinical Practice Guideline, the total serum bilirubin or transcutaneous bilirubin level should be measured in each infant in the first 24 hours of life. The concentration of bilirubin has been measured for 150 years. During that time the analytical methods for its determination have been significantly improved, the nomenclature of bilirubin has been also unified, but it is still unknown what concentration of bilirubin cause a life-threatening encephalopathy in the newborn. Under the current recommendations, clinical decisions to introduce phototherapy in the treatment of newborns’ hyperbilirubinemia are based on total bilirubin concentration, which is determined on biochemical analyzers and point of care testing systems. However, it is not always possible to predict encephalopathy based on the total bilirubin level. Probably in the future, as the availability of routine methods for the determination of unconjugated, free bilirubin becomes more available, measurement of “free” bilirubin will improve risk assessment for bilirubin neurotoxicity.

BACKGROUND:Orthodontic tooth movement (OTM) is a complex phenomenon mediated by cytokines, of which interleukin-1 beta (IL‑1β) is potently involved in the remodeling of the periodontal ligament (PDL) and bone. Whether the pattern of IL‑1β release differs at the sides of tension and compression is not yet clarified.OBJECTIVES:The aim of the present study was to evaluate the level of IL‑1β and the ratio of IL‑1β to interleukin-1 receptor antagonist (IL‑1RA) in gingival crevicular fluid (GCF) at the tension and compression sides during orthodontic canine retraction.MATERIAL AND METHODS:Seventeen patients scheduled for orthodontic treatment with bilateral extraction of maxillary first premolars and canine retraction were enrolled. Tooth 2.3 was retracted, teeth 1.3 and 3.3 served as controls. Gingival crevicular fluid samples were collected from the tension and compression sides of each tooth at baseline (before the 1st activation - day 0) and at days 2 and 7, and then again before the 2nd activation (day 28) and at days 30 and 35. The levels of IL‑1β and IL‑RA were evaluated with the enzyme-linked immunosorbent assay (ELISA).RESULTS:After the 1st activation, a statistically significant increase in the level of IL‑1β was observed at teeth 2.3 (p < 0.03 mesially and p < 0.05 distally) and 1.3 (p < 0.05 mesially and distally), both at the tension and compression sides. The 2nd activation resulted in a gradual increase in the IL‑1β level at both canines; however, statistical significance was reached only for tooth 2.3 (p < 0.05 mesially and p < 0.02 distally). In terms of the IL‑1β/IL‑1RA ratio, a significant increase was observed only at the compression side of the experimental tooth (p < 0.01).CONCLUSIONS:An increase in the IL‑1β level in GCF was observed both at the tension and compression sides of the actively retracted canine 2.3 as well as the contralateral canine 1.3; a significant rise in the IL‑1β/IL‑1RA ratio was noted only at the compression side of the experimental tooth 2.3, indicating the zone of active bone resorption.

Background Tryptophan metabolism via the kynurenine pathway is considered the link between the immune and endocrine systems. Dysregulation of serotonergic transmission can stem from the direct influence of interferon-α on the activity of serotonergic receptors 5-HT1A and 5-HT2A, and from its indirect effect on tryptophan metabolism. Induction of the kynurenine pathway increases the concentration of neurotoxic kynurenine metabolites, and the activity of kynurenine derivatives is linked to the onset of depression. The aim of our study was to evaluate the relationships between depressive symptoms and kynurenine, tryptophan, anthranilic acid and kynurenic acid concentrations, indolamine 2,3-dioxygenase (IDO) activity and tryptophan availability to the brain. Methods The study followed a prospective longitudinal cohort design. We evaluated 101 patients with chronic hepatitis C who were treated with pegylated interferon-α2a, and 40 controls who were awaiting treatment. We evaluated the relationships between total score on the Montgomery–Åsberg Depression Rating Scale and kynurenine, tryptophan, anthranilic acid and kynurenic acid concentrations, IDO activity and tryptophan availability to the brain. A logistic regression model was adapted for the diagnosis of major depressive disorder at each time point, taking into account changes in parameters of the kynurenine pathway between a given time point and the baseline measurement. Results Of the treated patients, 44% fulfilled the criteria for major depressive disorder at least once during the 24 weeks of treatment. Anthranilic acid concentrations were significantly increased compared to baseline for all time points except week 2. Tryptophan availability showed a significant decrease (β = −0.09, p = 0.01) only in week 12 of treatment. Over time, kynurenine, tryptophan and anthranilic acid concentrations, as well as IDO activity and tryptophan availability to the brain, were significantly associated with total score on the Montgomery–Åsberg Depression Rating Scale. A logistic regression model revealed that participants with decreased tryptophan availability to the brain at 12 weeks of treatment and participants with increased anthranilic acid concentrations at week 24 of treatment were at increased risk for diagnosis of major depressive disorder (odds ratios 2.92 and 3.59, respectively). Limitations This study had an open-label design in a population receiving naturalistic treatment. Conclusion The present study provides the first direct evidence of the role of anthranilic acid in the pathogenesis of inflammation-induced major depressive disorder during treatment for hepatitis C with pegylated interferon-α2a.

Introduction: The calcium-phosphate homeostasis is important for proper child growth and development and vitamin D plays a crucial role in this process. Aim: The aim of the study was to assess the relationship between total calcium (Ca), phosphates (Pi), parathyroid hormone (PTH) and the concentration of 25-hydroxycholecalciferol (25 (OH) D3) in children. Materials and methods: 164 serum samples were collected from children (1 day to 18 years) hospitalized in University Children’s Hospital in Krakow. Concentrations of Ca, Pi (dry chemistry-method), PTH (immunoradiometric-method) and 25(OH)D3 (HPLC) were determined. The results were analysed for all children, regardless of the age and for children under and above the age of four. Results: Analyzing all results, the mean of Ca concentration was significantly higher in the children with optimal 25(OH)D3 concentration compared to the children, who had 25(OH)D3 deficiency (p < 0.05). In samples from children ≤4 years of age, the significantly higher mean concentrations of Ca and Pi in the children with optimal 25(OH)D3 concentration compared to the children, who had suboptimal levels of 25(OH)D3 were found (p = 0.04; p = 0.002; respectively). For all results and for results from children ≤ 4 years of age, Ca and Pi concentrations were positively correlated with 25(OH)D3 concentration (p < 0.002, p < 0.01; p < 0.003, p < 0.004; respectively), there were no relationship between PTH and 25(OH)D3 concentrations. Conclusion: Comparing the results of calcium and phosphate concentration only to the appropriate reference ranges may not be sufficient to evaluate calcium-phosphate metabolism. Probably the reference values for total calcium and phosphates should be established depending on vitamin D levels, especially for young children.