Melanoma and Nevi Subtype Histopathological Characterization with Optical Coherence Tomography.

BACKGROUND: Melanoma incidence has continued to rise in the latest decades, and the forecast is not optimistic. Non-invasive diagnostic imaging techniques such as optical coherence tomography (OCT) are largely studied; however, there is still no agreement on its use for the diagnosis of melanoma. For dermatologists, the differentiation of non-invasive (junctional nevus, compound nevus, intradermal nevus, and melanoma in-situ) versus invasive (superficial spreading melanoma and nodular melanoma) lesions is the key issue in their daily routine. METHODS: This work performs a comparative analysis of OCT images using haematoxylin-eosin (HE) and anatomopathological features identified by a pathologist. Then, optical and textural properties are extracted from OCT images with the aim to identify subtle features that could potentially maximize the usefulness of the imaging technique in the identification of the lesion's potential invasiveness. RESULTS: Preliminary features reveal differences discriminating melanoma in-situ from superficial spreading melanoma and also between melanoma and nevus subtypes that pose a promising baseline for further research. CONCLUSIONS: Answering the final goal of diagnosing non-invasive versus invasive lesions with OCT does not seem feasible in the short term, but the obtained results demonstrate a step forward to achieve this.

Hospitalized population diagnosed with Covid-19 in public health centers in the southeastern region of Greater Buenos Aires / [Población hospitalizada con diagnóstico de Covid-19 en los centros de salud públicos de la región sudeste del Gran Buenos Aires]

Introduction: The present work describes the clinical characteristics and interventions to minimize morbidity and mortality in hospitalized patients diagnosed with COVID-19. Methods: It is a prospective cohort investigation of patients who received a response from the Health Centers in the southeast region (RS) of the metropolitan area (AMBA) from April 8 to September 30, 2020. A Situation Room was used epidemiological with two monitoring and follow-up boards, one for bed management and the other for patient management. Results: During the analyzed period, 2,588 patients with confirmed COVID-19 diagnosis were admitted, 1,943 with suspected COVID-19 pathology, and 1,464 subjects with other pathologies. 55% of the patients were men and the mean age was 51 years. There were 82.8% patients with pre-existing diseases, hypertension and diabetes were the most frequent. 14% were hospitalized in the Intensive Care Unit. The mortality of the cohort was 15.05%, mortality was higher for men, with a mean age of 60 years, 92.65% had some pre-existing disease. Conclusion: Our cohort is younger than other published works. Older people, men, and people with comorbidities are at increased risk for COVID-19-related mortality. The public health system was able to respond to the demand without collapsing the hospital institutions.

Introducción: En el presente trabajo se describen las características clínicas y las intervenciones para minimizar la morbimortalidad en pacientes hospitalizados con diagnóstico de COVID-19. Métodos: Es una investigación de cohorte prospectiva de pacientes que recibieron respuesta de los Centros de Salud en la región sudeste (RS) del área metropolitana (AMBA) desde el 8 de abril hasta el 30 de septiembre de 2020. Se utilizó una Sala de Situación epidemiológica con dos tableros de monitoreo y seguimiento, uno de gestión de camas y otro de gestión de pacientes. Resultados: Durante el periodo analizado se internaron2.588pacientes con diagnóstico COVID-19 confirmados, 1.943 con sospecha de patología COVID-19, y 1.464sujetos con otras patologías. El 55% de los pacientes eran hombres y la edad media fue de 51 años. Hubo 82,8% pacientes con enfermedades preexistentes, hipertensión y diabetes fueron las más frecuentes. El 14% fue hospitalizado en la Unidad de Terapia Intensiva. La mortalidad de la cohorte fue del 15,05%, la mortalidad fue mayor para los hombres, con una edad media de 60 años, el 92,65% tenía alguna enfermedad preexistente. Conclusión: Nuestra cohorte es más joven que otros trabajos publicados. Las personas mayores, los hombres y las personas con comorbilidades tienen mayor riesgo de mortalidad relacionada con COVID-19. El sistema de salud público pudo responder a la demanda sin llegar a colapsar las instituciones hospitalarias.

La región sudeste del Gran Buenos Aires frente a la pandemia Evaluación de la ampliación del sistema de salud en el marco de la COVID 19 / The Southeastern region of the Buenos Aires Province metropolitan area in the face of the pandemic: evaluation of the expansion of the health system in the framework of COVID-19

INTRODUCCIÓN: La región sudeste del Gran Buenos Aires (GBA) reformuló el sistema público de salud por la pandemia de COVID19. Entre las medidas que se tomaron, está la ampliación del número de camas mediante la construcción y puesta en marcha de tres hospitales. OBJETIVO: Evaluar el impacto de la ampliación del número de camas en los resultados de internación de los pacientes asistidos por los efectores públicos de salud durante el período de estudio (8 de abril de 2020 al 11 de septiembre de 2020). MÉTODOS: Estudio descriptivo a partir de información registrada en el Tablero COVID-19, software de gestión desarrollado por el equipo del Instituto del Cálculo de la Universidad de Buenos Aires, en el que se obtienen datos de cada paciente internado en la red de efectores de salud; se evalúan los resultados del efecto del aumento de la capacidad instalada. RESULTADOS: Se registraron 2 306 pacientes internados, de los cuales 266 (11,54%) requirieron internación en unidad de cuidados intensivos (UCO), 1 786 (77,4%) en cuidados intermedios y 254 (11%) pacientes en sala general. La media de edad fue de 50,63 y los pacientes de sexo masculino representaron el 55,5% del total. Se produjeron 253 muertes (10,97%), de las cuales el 64% fueron hombres. El 58,3% del total tenían enfermedades preexistentes, estos tienen un riesgo 90% más alto que quienes no las tenían. El promedio total de ocupación de camas en UCI fue del 40,7%, mientras que el de ocupación en cuidados intermedios fue de 61,5%. Sin los hospitales nuevos, 169 pacientes (9,46%) no hubieran tenido camas en cuidados intermedios y 31 pacientes (11,6%) no hubieran tenido cama en la UCI. DISCUSIÓN: El sistema de salud de la región sudeste del GBA se preparó de manera adecuada gracias a la ampliación del número de camas de internación.

Serum markers improve current prediction of metastasis development in early-stage melanoma patients: a machine learning-based study.

Metastasis development represents an important threat for melanoma patients, even when diagnosed at early stages and upon removal of the primary tumor. In this scenario, determination of prognostic biomarkers would be of great interest. Serum contains information about the general status of the organism and therefore represents a valuable source for biomarkers. Thus, we aimed to define serological biomarkers that could be used along with clinical and histopathological features of the disease to predict metastatic events on the early-stage population of patients. We previously demonstrated that in stage II melanoma patients, serum levels of dermcidin (DCD) were associated with metastatic progression. Based on the relevance of the immune response on the cancer progression and the recent association of DCD with local and systemic immune response against cancer cells, serum DCD was analyzed in a new cohort of patients along with interleukin 4 (IL-4), IL-6, IL-10, IL-17A, interferon γ (IFN-γ), transforming growth factor-ß (TGF- ß), and granulocyte-macrophage colony-stimulating factor (GM-CSF). We initially recruited 448 melanoma patients, 323 of whom were diagnosed as stages I-II according to AJCC. Levels of selected cytokines were determined by ELISA and Luminex, and obtained data were analyzed employing machine learning and Kaplan-Meier techniques to define an algorithm capable of accurately classifying early-stage melanoma patients with a high and low risk of developing metastasis. The results show that in early-stage melanoma patients, serum levels of the cytokines IL-4, GM-CSF, and DCD together with the Breslow thickness are those that best predict melanoma metastasis. Moreover, resulting algorithm represents a new tool to discriminate subjects with good prognosis from those with high risk for a future metastasis.

Influence of Lipid Fragmentation in the Data Analysis of Imaging Mass Spectrometry Experiments.

Imaging mass spectrometry (IMS) is becoming an essential technique in lipidomics. Still, many questions remain open, precluding it from achieving its full potential. Among them, identification of species directly from the tissue is of paramount importance. However, it is not an easy task, due to the abundance and variety of lipid species, their numerous fragmentation pathways, and the formation of a significant number of adducts, both with the matrix and with the cations present in the tissue. Here, we explore the fragmentation pathways of 17 lipid classes, demonstrating that in-source fragmentation hampers identification of some lipid species. Then, we analyze what type of adducts each class is more prone to form. Finally, we use that information together with data from on-tissue MS/MS and MS3 to refine the peak assignment in a real experiment over sections of human nevi, to demonstrate that statistical analysis of the data is significantly more robust if unwanted peaks due to fragmentation, matrix, and other species that only introduce noise in the analysis are excluded.

Influence of the Cation Adducts in the Analysis of Matrix-Assisted Laser Desorption Ionization Imaging Mass Spectrometry Data from Injury Models of Rat Spinal Cord.

Imaging mass spectrometry (IMS) is quickly becoming a technique of reference to visualize the lipid distribution in tissue sections. Still, many questions remain open, and data analysis has to be optimized to avoid interpretation pitfalls. Here we analyze how the variation on the [Na+]/[K+] relative abundance affects the detection of lipids between sections of spinal cord of (uninjured) control rats and of models of spinal cord demyelination and traumatic contusion injury. The [M + Na]+/[M + K]+ adducts ratio remained approximately constant along transversal and longitudinal sections of spinal cord from control animals, but it strongly changed depending on the type of lesion. A substantial increase in the abundance of [M + Na]+ adducts was observed in samples from spinal cord with demyelination, while the intensity of the [M + K]+ adducts was stronger in those sections from mechanically injured spinal cords. Such changes masked the modifications in the lipid profile due to the injury and only after summing the signal intensity of all adducts and corresponding monoprotonated molecular ions of each detected lipid in a single variable, it was possible to unveil the real changes in the lipid profile due to the lesion. Such lipids included glycerophospholipids (both diacyl and aryl-acyl), sphingolipids, and nonpolar lipids (diacyl and triacylglycerols), which are the main lipid classes detected in positive-ion mode. Furthermore, the results demonstrate the sensitivity of the technique toward modification in tissue homeostasis and that the [M + Na]+/[M + K]+ ratio may be used to detect alterations in such homeostasis.

Lipid fingerprint image accurately conveys human colon cell pathophysiologic state: A solid candidate as biomarker.

Membrane lipids are gaining increasing attention in the clinical biomarker field, as they are associated with different pathologic processes such as cancer or neurodegenerative diseases. Analyzing human colonoscopic sections by matrix assisted laser/desorption ionization (MALDI) mass spectrometry imaging techniques, we identified a defined number of lipid species changing concomitant to the colonocyte differentiation and according to a quite simple mathematical expression. These species felt into two lipid families tightly associated in signaling: phosphatidylinositols and arachidonic acid-containing lipids. On the other hand, an opposed pattern was observed in lamina propria for AA-containing lipids, coinciding with the physiological distribution of the immunological response cells in this tissue. Importantly, the lipid gradient was accompanied by a gradient in expression of enzymes involved in lipid mobilization. Finally, both lipid and protein gradients were lost in adenomatous polyps. The latter allowed us to assess how different a single lipid species is handled in a pathological context depending on the cell type. The strict patterns of distribution in lipid species and lipid enzymes described here unveil the existence of fine regulatory mechanisms orchestrating the lipidome according to the physiological state of the cell. In addition, these results provide solid evidence that the cell lipid fingerprint image can be used to predict precisely the physiological and pathological status of a cell, reinforcing its translational impact in clinical research.

Deciphering the Lipid Architecture of the Rat Sciatic Nerve Using Imaging Mass Spectrometry.

Knowledge on the normal structure and molecular composition of the peripheral nerves is essential to understand their pathophysiology and to select the regeneration strategies after injury. However, the precise lipid composition of the normal peripheral nerve is still poorly known. Here, we present the first study of distribution of individual lipids in the mature sciatic nerve of rats by imaging mass spectrometry. Both positive and negative ion modes were used to detect, identify and in situ map 166 molecular species of mainly glycerophospholipids, sphingomyelins, sulfatides, and diacyl and triacylglycerols. In parallel, lipid extracts were analyzed by LC-MS/MS to verify and complement the identification of lipids directly from the whole tissue. Three anatomical regions were clearly identified by its differential lipid composition: the nerve fibers, the connective tissue and the adipose tissue that surrounds the nerve. Unexpectedly, very little variety of phosphatidylcholine (PC) species was found, being by far PC 34:1 the most abundant species. Also, a rich composition on sulfatides was detected in fibers, probably due to the important role they play in the myelin cover around axons, as well as an abundance of storage lipids in the adipose and connective tissues. The database of lipids here presented for each region and for the whole sciatic nerve is a first step toward understanding the variety of the peripheral nerves' lipidome and its changes associated with different diseases and mechanical injuries.

Identification of Biomarkers of Necrosis in Xenografts Using Imaging Mass Spectrometry.

Xenografts are commonly used to test the effect of new drugs on human cancer. However, because of their heterogeneity, analysis of the results is often controversial. Part of the problem originates in the existence of tumor cells at different metabolic stages: from metastatic to necrotic cells, as it happens in real tumors. Imaging mass spectrometry is an excellent solution for the analysis of the results as it yields detailed information not only on the composition of the tissue but also on the distribution of the biomolecules within the tissue. Here, we use imaging mass spectrometry to determine the distribution of phosphatidylcholine (PC), phosphatidylethanolamine (PE), and their plasmanyl- and plasmenylether derivatives (PC-P/O and PE-P/O) in xenografts of five different tumor cell lines: A-549, NCI-H1975, BX-PC3, HT29, and U-87 MG. The results demonstrate that the necrotic areas showed a higher abundance of Na(+) adducts and of PC-P/O species, whereas a large abundance of PE-P/O species was found in all the xenografts. Thus, the PC/PC-ether and Na(+)/K(+) ratios may highlight the necrotic areas while an increase on the number of PE-ether species may be pointing to the existence of viable tumor tissues. Furthermore, the existence of important changes in the concentration of Na(+) and K(+) adducts between different tissues has to be taken into account while interpreting the imaging mass spectrometry results. Graphical Abstract ᅟ.

Optimized Protocol To Analyze Changes in the Lipidome of Xenografts after Treatment with 2-Hydroxyoleic Acid.

Xenografts are a popular model for the study of the action of new antitumor drugs. However, xenografts are highly heterogeneous structures, and therefore it is sometimes difficult to evaluate the effects of the compounds on tumor metabolism. In this context, imaging mass spectrometry (IMS) may yield the required information, due to its inherent characteristics of sensitivity and spatial resolution. To the best of our knowledge, there is still no clear analysis protocol to properly evaluate the changes between samples due to the treatment. Here we present a protocol for the evaluation of the effect of 2-hydroxyoleic acid (2-OHOA), an antitumor compound, on xenografts lipidome based on IMS. Direct treated/control comparison did not show conclusive results. As we will demonstrate, a more sophisticated protocol was required to evaluate these changes including the following: (1) identification of different areas in the xenograft, (2) classification of these areas (necrotic/viable) to compare similar types of tissues, (3) suppression of the effect of the variation of adduct formation between samples, and (4) normalization of the variables using the standard deviation to eliminate the excessive impact of the stronger peaks in the statistical analysis. In this way, the 36 lipid species that experienced the largest changes between treated and control were identified. Furthermore, incorporation of 2-hydroxyoleic acid to a sphinganine base was also confirmed by MS/MS. Comparison of the changes observed here with previous results obtained with different techniques demonstrates the validity of the protocol.

Imaging mass spectrometry increased resolution using 2-mercaptobenzothiazole and 2,5-diaminonaphtalene matrices: application to lipid distribution in human colon.

Imaging mass spectrometry is becoming a reference technique in the field of lipidomics, due to its ability to map the distribution of hundreds of species in a single run, along a tissue section. The next frontier is now achieving increasing resolution powers to offer cellular (or even sub-cellular) resolution. Thus, the new spectrometers are equipped with sophisticated optical systems to decrease the laser spot to <30 µm. Here, we demonstrate that by using the correct matrix (i.e., a matrix that maximizes ion detection and forms small crystals) and a careful preparation, it is possible to achieve resolutions of ∼5-10 µm, even with spectrometers equipped with non-optimal optics, which produces laser spots of 50 µm or even larger. As a proof of concept, we present images of distributions of lipids, both in positive and negative ion mode, over human colon endoscopic sections, recorded using 2-mercaptobenzothiazole for positive ion mode and 2,5-diaminonaphtalene for negative ion mode and an LTQ-Orbitrap XL, equipped with a matrix-assisted laser desorption ionization (MALDI) source that produces astigmatic laser spots. Graphical Abstract Imaging mass spectrometry is becoming an invaluable technique to complement traditional histology, but still higher resolutions are required. Here we deal with such issue.

6R-tetrahydrobiopterin treated PKU patients below 4 years of age: Physical outcomes, nutrition and genotype.

BACKGROUND AND AIMS: Phenylalanine-restricted diets have proven effective in treating phenylketonuria. However, such diets have occasionally been reported to hinder normal development. Our study aimed to assess whether treating 0-4-year-old phenylketonuric patients with 6R-tetrahydrobiopterin might prevent growth retardation later in life. METHODS: We conducted a longitudinal retrospective study which examined anthropometric characteristics of phenylketonuric patients on 6R-tetrahydrobiopterin therapy (22 subjects), and compared them with a group of phenylketonuric patients on protein-restricted diets (44 subjects). Nutritional issues were also considered. We further explored possible relationships between mutations in the PAH gene, BH4 responsiveness and growth outcome. RESULTS: No significant growth improvements were observed in either the group on 6R-tetrahydrobiopterin treatment (height Z-score: initial= -0.57 ± 1.54; final=-0.52 ± 1.29; BMI Z-score: initial=0.17 ± 1.05; final=0.18 ± 1.00) or the diet-only group (height Z-score: initial=-0.92 ± 0.96; final= -0.78 ± 1.08; BMI Z-score: initial=0.17 ± 0.97; final=-0.07 ± 1.03) over the 1-year observation period. Furthermore, we found no significant differences (p>0.05) between the two groups at any of the time points considered (0, 6 and 12 months). Patients on 6R-tetrahydrobiopterin increased their phenylalanine intake (from 49.1 [25.6-60.3] to 56.5 [39.8-68.3] mgkg(-1)day(-1)) and natural protein intake (from 1.0 [0.8-1.7] to 1.5 [1.0-1.8] g kg(-1)day(-1)), and some patients managed to adopt normal diets. Higher phenylalanine and natural protein intakes were positively correlated with better physical outcomes in the diet-only group (p<0.05). No correlation was found between patient genotype and physical outcomes, results being similar regardless of the nutritional approach used. We did not detect any side effects due to 6R-tetrahydrobiopterin administration. CONCLUSIONS: Our study indicates that treating 0-4-year-old phenylketonuric patients with 6R-tetrahydrobiopterin is safe. However, poor developmental outcomes were observed, despite increasing the intake of natural proteins. Genotype could be a valid predictor of tetrahydrobiopterin-responsiveness, since patients who carried the same genotype responded similarly to the 6R-tetrahydrobiopterin loading test. On the other hand, harbouring 6R-tetrahydrobiopterin responsive genotypes did not predispose patients to better physical outcomes.

Quantification of arginine and its methylated derivatives in healthy children by liquid chromatography-tandem mass spectrometry.

Asymmetric dimethylarginine (ADMA) is a competitive inhibitor of nitric oxide synthase, which is responsible for most of the vascular nitric oxide (NO) produced. NO is an important physiological mediator of vascular tone and structure in normally functioning endothelial cells. We report the optimization of a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for simultaneous determination of arginine (Arg) and its derivatives in biological samples. Chromatographic separation and mass detection were performed by reverse phase chromatography coupled with tandem mass spectrometry. For sample preparation, plasma proteins were removed by centrifugal filters. Positive electrospray ionization was performed and analytes were detected by multiple reaction monitoring. Inter- and intra-day repeatability, accuracy, recovery, and limits of detection and quantification were evaluated to validate the method. Plasma and urine levels were measured in healthy children to establish control values: 52.2-124.7 µM for Arg, 0.06-0.16 µM for MMA, 0.42-1.10 µM for ADMA and 0.41-0.96 µM for SDMA in plasma. Quantification of Arg and its methylated derivatives by LC-MS/MS can be carried out without the need of organic solvents for sample preparation, and be used as a valuable tool in research on endothelial dysfunction.

Analysis of the lipidome of xenografts using MALDI-IMS and UHPLC-ESI-QTOF.

Human tumor xenografts in immunodeficient mice are a very popular model to study the development of cancer and to test new drug candidates. Among the parameters analyzed are the variations in the lipid composition, as they are good indicators of changes in the cellular metabolism. Here, we present a study on the distribution of lipids in xenografts of NCI-H1975 human lung cancer cells, using MALDI imaging mass spectrometry and UHPLC-ESI-QTOF. The identification of lipids directly from the tissue by MALDI was aided by the comparison with identification using ESI ionization in lipid extracts from the same xenografts. Lipids belonging to PCs, PIs, SMs, DAG, TAG, PS, PA, and PG classes were identified and their distribution over the xenograft was determined. Three areas were identified in the xenograft, corresponding to cells in different metabolic stages and to a layer of adipose tissue that covers the xenograft.

Anthropometric characteristics and nutrition in a cohort of PAH-deficient patients.

BACKGROUND & AIMS: Treating phenylketonuria based upon strict vegetarian diets has occasionally been found to hamper physical development, some patients presenting with growth retardation and malnutrition. In addition, some researchers have reported an association between higher protein intakes and attaining better developmental outcomes, although it remains unclear which protein fraction (natural or synthetic) has the greatest influence on growth. The present study aimed to evaluate anthropometric characteristics and nutrition in a cohort of patients with phenylketonuria and mild-hyperphenylalaninaemia from birth to adulthood. METHODS: We conducted a retrospective longitudinal study comparing anthropometric characteristics (weight, height, body mass index, and growth rate) in our patients and healthy subjects, with the measurements expressed as z-scores. Nutritional issues were also considered. Data were collected every 6 months from birth to 18 years of age. RESULTS: Growth impairment was observed in phenylketonuric patients. Specifically, there were two well-differentiated periods throughout which height fell well below z-score = 0: from birth to two years of age, and on reaching adulthood. We also found height and weight to be positively correlated with phenylalanine intake. No growth retardation was seen in the patients with mild-hyperphenylalaninaemia. CONCLUSIONS: Phenylketonuric patients showed growth impairment in the early stages, with higher phenylalanine intakes being associated with attaining better developmental outcomes in this period. Therefore, prescribing very stringent diets in the early years might predispose phenylketonuric patients to retarded growth later in life, with growth outcomes in adulthood being well below the 50th percentile for healthy subjects.

Tetrahydrobiopterin therapy vs phenylalanine-restricted diet: impact on growth in PKU.

BACKGROUND: Treatment of phenylketonuria based upon strict vegetarian diets, with very low phenylalanine intake and supplemented by phenylalanine-free formula, has proven to be effective in preventing the development of long-term neurological sequelae due to phenylalanine accumulation. On the other hand, such diets have occasionally been reported to hinder normal development, some individuals presenting with growth retardation. Tetrahydrobiopterin therapy has opened up new treatment options for a significant proportion of phenylketonuric patients, enabling them to eat normal diets and be freed from the need to take synthetic supplements. However, little is known about how this therapy affects their physical development. METHODS: We conducted a retrospective longitudinal study examining anthropometric characteristics (height, weight, body mass index and growth speed Z-scores) in a cohort of phenylketonuric patients on tetrahydrobiopterin therapy (38 subjects) comparing their characteristics with those of a group of phenylketonuric patients on phenylalanine-restricted diets (76 subjects). Nutritional issues were also considered, to further explore the possibility of higher natural protein intake being associated with better physical development. Data were collected every six months over two different periods of time (two or five years). RESULTS: No improvement was observed in the aforementioned anthropometric variables in the cohort on tetrahydrobiopterin therapy, from prior to starting treatment to when they had been taking the drug for two or five years. Rather, in almost all cases there was a fall in the mean Z-score for the variables during these periods, although the changes were not significant in any case. Further, we found no statistically differences between the two groups at any considered time point. Growth impairment was also noted in the phenylketonuric patients on low-phenylalanine diets. Individuals on tetrahydrobiopterin therapy increased their natural protein intake and, in some instances, this treatment enabled individuals to eat normal diets, with protein intake meeting RDAs. No association was found, however, between higher protein intake and growth. CONCLUSION: Our study identified growth impairment in patients with phenylketonuria on tetrahydrobiopterin, despite higher intakes of natural proteins. In fact, individuals undergoing long-term tetrahydrobiopterin treatment seemed to achieve similar developmental outcomes to those attained by individuals on more restricted diets.

Arginine-guanidinoacetate-creatine pathway in preterm newborns: creatine biosynthesis in newborns.

The phosphocreatine/creatine system is fundamental for the proper development of the embryonic brain. Being born prematurely might alter the creatine biosynthesis pathway, in turn affecting creatine supply to the developing brain. We enrolled 53 preterm and very preterm infants and 55 full-term newborns. The levels of urinary guanidinoacetate, creatine, creatinine and amino acids were measured in the preterm and very preterm groups, 48 h and 9 days after birth and at discharge, and 48 h after birth in the full-term group. Guanidinoacetate concentrations of both preterm and very preterm newborns were significantly higher at discharge than the values for the full-term group at 48 h, while very preterm infants showed urinary creatine values significantly lower than those measured in the full-term group. Our results suggest an impairment of the creatine biosynthesis pathway in preterm and very preterm newborns, which could lead to creatine depletion affecting the neurological outcome in prematurely born infants.

New evidence for assessing tetrahydrobiopterin (BH(4)) responsiveness.

OBJECTIVE: To evaluate the protocol we propose for detecting BH(4)-responsive patients and the possibility of delimiting more precisely the population to be tested. METHODS: We recruited 102 phenylketonuric patients on a phenylalanine (Phe)-restricted diet. The initial stage of the protocol was a 24-h BH(4) loading test involving Phe loading and subsequent ingestion of the cofactor, a 50% fall in blood Phe levels being considered a positive response. The non-responders at this stage then completed a one-week therapeutic test combining BH(4) administration and daily protein intake meeting recommended dietary allowances, to assess whether the 24-h test had detected all responders. RESULTS: The 24-h test detected almost all BH(4) responders (30.3% of the 99 patients included in the analysis), with just two patients (2.0%) subsequently responding positively to the therapeutic test. The 24-h test did not give any false positive results. CONCLUSIONS: The 24-h BH(4) loading test is clinically useful for screening phenylketonuric patients. Specifically, 95% of patients with Phe levels <700 µmol/L, and none with Phe levels >1500 µmol/L were BH(4)-responsive. Given these results, we conclude that patients with Phe levels<700 µmol/L or>1500 µmol/L probably do not need to be tested, prioritising the identification of BH(4)-responsiveness among individuals with intermediate Phe concentrations, between the aforementioned values. Additionally, our results suggest that the therapeutic test only needs to be performed in cases where the reduction in blood Phe levels after cofactor administration is within the range 40%-50%.