Abnormal concentrations of acetylated amino acids in cerebrospinal fluid in acetyl-CoA transporter deficiency.

Acetyl-CoA transporter 1 (AT-1) is a transmembrane protein which regulates influx of acetyl-CoA from the cytosol to the lumen of the endoplasmic reticulum and is therefore important for the posttranslational modification of numerous proteins. Pathological variants in the SLC33A1 gene coding for AT-1 have been linked to a disorder called Huppke-Brendel syndrome, which is characterized by congenital cataracts, hearing loss, severe developmental delay and early death. It has been described in eight patients so far, who all had the abovementioned symptoms together with low serum copper and ceruloplasmin concentrations. The link between AT-1 and low ceruloplasmin concentrations is not clear, nor is the complex pathogenesis of the disease. Here we describe a further case of Huppke-Brendel syndrome with a novel and truncating homozygous gene variant and provide novel biochemical data on N-acetylated amino acids in cerebrospinal fluid (CSF) and plasma. Our results indicate that decreased levels of many N-acetylated amino acids in CSF are a typical metabolic fingerprint for AT-1 deficiency and are potential biomarkers for the defect. As acetyl-CoA is an important substrate for protein acetylation, we performed N-terminal proteomics, but found only minor effects on this particular protein modification. The acetyl-CoA content in patient's fibroblasts was insignificantly decreased. Our data may help to better understand the mechanisms underlying the metabolic disturbances, the pathophysiology and the clinical phenotype of the disease.

HOW TO TREAT PATIENTS AFTER SERIOUS ADVERSE EFFECTS CAUSED BY TNF INHIBITORS?

Biological agents are widely used in the treatment of autoimmune rheumatic disorders. We report on serious adverse events during treatment with anti-tumor necrosis factor antibody in two of our patients with juvenile idiopathic arthritis. One patient was treated with a biological agent due to juvenile idiopathic arthritis complicated by uveitis, developing miliary tuberculosis during treatment. After treatment with antituberculotics, she recovered completely. Her underlying disease is currently in remission. Another patient was treated for juvenile spondyloarthritis and developed an inflammatory process of the central nervous system with serious neurological deficits. He was treated with high-dose corticosteroids, followed by slowly tapering doses of corticosteroids. His neurological deficits improved, but are still present. Similar cases have been described previously, but there are no recommendations how to treat arthritis afterwards in such patients. We would like to emphasize the need of developing guidelines for further treatment of arthritis after the occurrence of serious adverse effects during treatment with biological agents.

Daytime Sleepiness from Preschool Children's and Parents' Perspectives: Is There a Difference?

This cross-sectional study investigated the level of daytime sleepiness and sleep-related behaviors in preschool children and compared their self-evaluations with the evaluations of their parents. It was conducted in Split-Dalmatian County, Croatia, among 196 preschool children aged 6-7 years seen at regular medical examinations, accompanied by their parents, using the Epworth sleepiness scale for children and parents/caregivers. Compared to their child's reports, parents tended to underestimate their child's sleepiness while sitting in a classroom at school (p = 0.001) and overestimate their child's sleepiness when lying down to rest or nap in the afternoon (p < 0.001). Boys were sleepier while sitting in a classroom at school during the morning than girls (p = 0.032). As much as 48.2% of preschool children had their own cellphones/tablets. Boys used video games (p < 0.001) and cellphones/tablets more than girls did (p = 0.064). Parental estimation of children playing video games at bedtime was lower than the child's report (p < 0.001). Children who had a TV in their bedroom reported more daytime sleepiness (p = 0.049), and those who played video games at bedtime went to sleep later during the weekend (p = 0.024). Also, children owning cellphone/tablets had longer sleep latency during the weekend compared to children not owning a cellphone (p = 0.015). This study confirmed that parents tend to underestimate children's habits of playing video games at bedtime and children's sleepiness during morning classes. Preschool children who use electronic devices at bedtime more frequently have prolonged sleep latency. These findings provide further evidence of the effects of electronic media devices on preschoolers' sleep patterns and daytime sleepiness.

Association of Appendicitis, Helicobacter Pylori Positive Gastritis and Thrombotic Thrombocytopenic Purpura in an Adolescent.

BACKGROUND Thrombotic thrombocytopenic purpura (TTP) in children is a rare life-threatening syndrome, characterized by microangiopathic hemolytic anemia, thrombocytopenia with renal dysfunction, neurologic symptoms, and fever. TTP is usually caused by deficient activity of von Willebrand factor cleaving protease (ADAMTS13), due to either gene mutations or acquired via anti-ADAMTS13 autoantibodies. It can be triggered by bone marrow or solid organ transplantation, cardiothoracic-, abdominal-, and orthopedic surgeries, infections including very rarely Helicobacter pylori infection. CASE REPORT Here we report a case of a 16-year-old male with TTP, who presented with thrombocytopenia before an appendectomy. Seven days after surgery, our patient started to vomit, developed melena, and was admitted to our pediatric intensive care unit (PICU) with clinical presentation of shock. Gastroscopy revealed H. pylori positive hemorrhagic gastritis. The patient was treated by erythrocyte transfusions, fresh frozen plasma, human albumin, glucose-electrolyte solutions, vitamin K, platelet transfusion before implantation of central venous catheter, and antibiotics. After 36 hours, we started plasma exchange (PEX). Blood tests showed deficiency of ADAMTS13. Due to the presence of anti-ADAMTS13 autoantibodies, rituximab was administered. Due to generalized tonic-clonic seizures, he was artificially ventilated. Brain MR angiography showed small ischemic cerebro-vascular insult in the arteria cerebri media region. Despite immunosuppressive therapy and PEX, the patient did not improve completely until the H. pylori infection was eradicated. After which, he recovered completely. CONCLUSIONS We present a rare case of TTP accompanied with appendicitis and gastritis caused by H. pylori, where TTP improvement was dependent on H. pylori infection eradication.

Functional outcome of children treated in intensive care unit.

OBJECTIVE: Outcome of patients is determined not only by severity of illness index, but also by the impact of patients' preadmission comorbid status. Therefore, we aimed at evaluating the outcome of patients treated in a pediatric intensive care unit, with special focus on the group of children with chronic diseases. METHODS: Data were obtained prospectively and outcome was assessed according to the Pediatric Overall Performance Category scale for 449 patients in a pediatric intensive care unit of the Split University Hospital. Functional performance was assessed as the preadmission score and the discharge score in patients with neurodevelopmental disabilities, patients with other chronic diseases, and those without chronic disease. RESULTS: The discharge functional status was significantly dependent on the preadmission functional status and on predicted mortality. Children with neurodevelopmental disabilities had the significantly worse baseline score and the significantly smaller deterioration of functional morbidity at discharge compared to children with no chronic disease and children with other chronic diseases. CONCLUSIONS: The Pediatric Overall Performance Category scale has proved its applicability in a small intensive care unit, with a heterogeneous population of patients. It should therefore be considered for regular evaluation of health care quality, as a simple and accurate tool. As opposed to other patients, functional status of children with neurodevelopmental disabilities was markedly influenced by their comorbidity. Their preadmission status was worse than the status of other children, and hence could not significantly deteriorate at discharge.

Formula for the prediction of apnea / hypopnea index in children with obstructive sleep apnea without polysomnography according to the clinical parameters: Is it reliable?

PURPOSE OF THE STUDY: The aim of the study was to propose "the risk formula" for obstructive sleep apnea in children according to the general and local clinical parameters and findings relevant for obstructive sleep apnea (OSA) severity. The unmet need for this formula arises from the economic burden of polysomnography (device, staff, training, special sleep centers, etc) as the golden standard for the diagnostics. MATERIALS AND METHODS USED: The study was performed from January 2013 until January 2016 in the Sleep Center, Department for Neuroscience, School of Medicine of the University of Split, Department of Pediatrics, University Hospital Split, Croatia and ENT Dept. University Hospital in Split, Croatia. Inclusion criteria were: age > two years, AHI >1 diagnosed by polysomnography. Exclusion criteria were: chronic lung disease, active tonsillitis/pharyngitis at the time of the physical exam and syndromes that affect breathing. All polysomnograms were scored by a qualified sleep technologist and interpreted by two board certified sleep physicians independently. Age, sex, BMI, Mallampati score, tonsillar size and adenoids size were recorded. All statistical calculations were performed using SPSS 20. RESULTS: In total 60 children were included in the study. The median of age was 5 years (range 2-9). There were 19 (32%) girls and 41 (68%) boys. Of all evaluated predictors, there were statistically significant differences in the values of AHI among children with different modified Mallampati score (χ2 = 28.2; p < 0.001), different size of tonsils (χ2 = 25.3; p < 0.001) and different size of adenoids (z = 2.7; p = 0,006) in univariate regression analysis. Strong positive association of AHI with modified Mallampati score (standardized B = 0.51; partial correlation = 0.542, r = 0.631) was found, as well as positive correlation of AHI with tonsillar size (standardized B = 0.246; partial correlation = 0.295,R = 0.489) in the multivariate forward stepwise regression analysis. CONCLUSION: Even though we are aware that PSG is the gold standard for diagnostics of SDB there is a significant financial burden for this diagnostic procedure. That is why there is a necessity for establishing good clinical standards and possible formula for OSA severity evaluation. We propose formula which includes Mallampati score and tonsillar size for OSA -risk calculation in order to perform early therapeutic intervention thereby reducing the risk of long-term negative consequences. We recommend this formula as the screening formula in circumstances where PSG is not available, in cases when the "waiting list" is too long or when a child can not cooperate to perform it. In developing countries like Croatia on time intervention with reduced procedure-associated costs is of the utmost importance.

High incidence of glucose-6-phosphate dehydrogenase deficiency in Croatian island isolate: example from Vis island, Croatia.

AIM: To determine the prevalence of glucose-6-phosphate dehydrogenase (G6PD) deficiency in the population of the town of Komiza on the island of Vis, which has previously been reported as a place with several cases of favism. METHODS: We screened 302 randomly selected men, using the fluorescent spot test. Fluorescence readings were performed at the beginning and 5, 10, and 20 minutes after incubation, and were classified into three groups: bright fluorescence, weak fluorescence, and no fluorescence. All men found to be G6PD deficient were tested with a quantitative spectrophotometric UV method. RESULTS: Of the 302 tested blood samples, 36 (11.9%) samples showed weak fluorescence or no fluorescence spots. Spectrophotometric UV test showed that 18 (5.96%) men were G6PD deficient. The prevalence of G6PD deficiency in the population of Komiza is significantly higher (P<0.001) than the prevalence in the whole population of Dalmatia in the south of Croatia (0.75% in men). CONCLUSION: On the basis of these findings, we recommend including the newborns from the island of Vis into a screening program for G6PD deficiency. Our results indicate that G6PD deficiency should be determined for all the island isolates in the Mediterranean basin and they warrant further studies.

Influence of the Inherited Glucose-6-phosphate Dehydrogenase Deficiency on the Appearance of Neonatal Hyperbilirubinemia in Southern Croatia.

BACKGROUND: Neonatal hyperbilirubinemia is a common clinical manifestation of the inherited glucose-6-phosphate dehydrogenase (G6PD) deficiency. AIM OF THE STUDY: The aim of this study was to investigate the influence of the inherited G6PD deficiency on the appearance of neonatal hyperbilirubinemia in southern Croatia. METHODS: The fluorescent spot test (FST) was used in a retrospective study to screen blood samples of 513 male children who had neonatal hyperbilirubinemia, of unknown cause, higher than 240 µmol/L. Fluorescence readings were performed at the beginning and at the fifth and tenth minute of incubation and were classified into three groups bright fluorescence (BF), weak fluorescence (WF) and no fluorescence (NF). Normal samples show bright fluorescence. All NF and WF samples at the fifth minute were quantitatively measured using the spectrophotometric method. RESULTS: Bright fluorescence was present in 461 patients (89.9%) at the fifth minute. The remaining 52 (10.1%) were quantitatively estimated using the spectrophotometric method. G6PD deficiency was observed in 38 patients (7.4%). CONCLUSIONS: Prevalence rate of G6PD deficiency among male newborns with hyperbilirubinemia in southern Croatia is significantly higher (p < 0.01) compared with the previously reported prevalence rate among male in general population of southern Croatia (0.75%). We recommend FST to be performed in hyperbilirubinemic newborns in southern Croatia.

Prevalence of factor V Leiden and G6PD 1311 silent mutations in Dalmatian population.

BACKGROUND: Factor V Leiden has been described as a common genetic risk factor for venous thromboembolism. The geographic distribution of this abnormality varies greatly, being high in Europe and almost absent in Asia and Africa. Particularly high prevalence is observed in some Mediterranean countries, which suggests the Mediterranean origin of this mutation. Similarly, prevalence of silent mutation 1311 of the G6PD gene seems to be higher among Mediterranean populations. Since the Dalmatian population (of south Croatia) geographically belongs to the Mediterranean populations we analyzed the prevalence of FV-Leiden and silent mutation 1311 in this region. Furthermore, because the coincidence of G6PD deficiency and venous thromboembolism was described earlier, we tested a possible association of FV-Leiden and G6PD deficiency. METHODS: One hundred sixty-eight healthy blood donors and 55 G6PD deficient individuals originating from the Dalmatian region were tested for the presence of FV-Leiden mutation and silent mutation 1311. RESULTS: Prevalence of FV-Leiden among blood donors was 2.4%, while among G6PD deficient individuals it was significantly higher, 11% (p=0.011). Prevalence of silent mutation 1311 among blood donors and G6PD deficient individuals was 21 and 15%, respectively. CONCLUSIONS: Observed allele frequencies among individuals originating from the Dalmatian region is similar to the neighboring European and Mediterranean populations. Interestingly, our results indicate the association of the FV-Leiden and G6PD deficiency and warrant further studies.

Immune Modulation Therapy in a CRIM-Positive and IgG Antibody-Positive Infant with Pompe Disease Treated with Alglucosidase Alfa: A Case Report.

Pompe disease is characterized by deficiency or absence of activity of the lysosomal enzyme acid alpha-glucosidase. As a result of ineffective metabolism, glycogen progressively accumulates in muscle tissues. Patients with an aggressive classic infantile-onset form generally rapidly die of cardiorespiratory failure. A cross-reactive immunological material (CRIM)-negative status is predictive of high anti-alglucosidase alfa antibody titers and usually a poor clinical outcome of enzyme replacement therapy (ERT). CRIM-positive patients can also develop robust antibody titers complicating therapeutic management.We successfully used an immune modulation therapy (IMT) protocol in a CRIM-positive infantile-onset patient with Pompe disease in whom infusions had to be temporarily discontinued because of safety concerns despite administration of pre-infusion medication. Prior to discontinuation, she had shown signs of clinical deterioration and continuous ventilation support through a tracheostomy was required. She was found to be positive for anti-alglucosidase alfa antibodies (1:6,400). IMT (rituximab, methotrexate and intravenous gamma globulin) was started, ERT was safely reintroduced during the IMT induction phase and, subsequently, the enzyme dose was increased, all without any complications. Antibodies disappeared, IMT was tapered and discontinued, and cadiomyopathy steadily improved. During 1 year of follow-up, she remained ventilator dependent and no gains in motor skills were noticed; motor functions will be closely monitored during sustained ERT.Although the reversal of clinical decline in our CRIM-positive and antibody-positive infant with Pompe disease cannot be solely attributed to IMT, our experiences with this protocol may be helpful to other physicians encountering comparable therapeutic dilemmas.

Characterization of G6PD deficiency in southern Croatia: description of a new variant, G6PD Split.

Glucose-6-phosphate dehydrogenase (G6PD) deficiency protects from severe forms of malaria. It is interesting therefore to analyze the molecular basis underlying G6PD deficiency in regions such as the Mediterranean basin where malaria was present for a long time in history. Here we report on the genetic characterization of G6PD deficiency among inhabitants of one Mediterranean region-the Dalmatian region of south Croatia. We analyzed 24 unrelated G6PD-deficient male subjects. Molecular testing revealed several different mutations: G6PD Cosenza 9, G6PD Mediterranean 4, G6PD Seattle 3, G6PD Union 3, and G6PD Cassano 1. Furthermore, we have identified one novel G6PD variant that we named G6PD Split. This variant is caused by a nucleotide change 1442 C-->G leading to the amino acid substitution 481 Pro-->Arg and is characterized by moderate enzyme deficiency (class III variant). This study reveals a higher prevalence (37.5%) of the Cosenza mutation in the Dalmatian region than anywhere else previously investigated and overall shows the considerable molecular heterogeneity underlining G6PD deficiency that can be observed in Mediterranean populations.

Desfecho funcional de crianças tratadas em unidade de terapia intensiva / Functional outcome of children treated in intensive care unit

OBJETIVO: O desfecho de pacientes não é somente determinado pelo índice de gravidade de doença, mas também pelo impacto do estado pré-admissão de comorbidade dos pacientes. Portanto, este artigo buscou avaliar o desfecho de pacientes tratados em uma unidade de terapia intensiva pediátrica, com foco especial no grupo de crianças com doenças crônicas. MÉTODOS: Os dados foram obtidos prospectivamente, e o desfecho foi avaliado segundo a escala Pediatric Overall Performance Category para 449 pacientes de uma unidade de terapia intensiva pediátrica do Split University Hospital. O desempenho funcional foi avaliado como o escore pré-admissão e o escore na alta hospitalar em pacientes com alterações neurodesenvolvimentais, com outras doenças crônicas e sem doença crônica. RESULTADOS: O estado funcional à alta hospitalar foi significativamente dependente do estado funcional pré-admissão e da mortalidade prevista. Crianças com alterações neurodesenvolvimentais apresentaram escore basal significativamente pior e deterioração de morbidade funcional na alta hospitalar significativamente menor, comparadas com crianças sem doença crônica e com crianças com outras doenças crônicas. CONCLUSÕES: A escala Pediatric Overall Performance Category demonstrou sua aplicabilidade em uma pequena unidade de terapia intensiva com uma população heterogênea de pacientes. Deve, portanto, ser considerada para avaliação regular de qualidade de cuidados à saúde como uma ferramenta simples e precisa. Ao contrário do que acontece com outros pacientes, o estado funcional de crianças com alterações neurodesenvolvimentais foi marcadamente influenciado por sua comorbidade. Seu estado pré-admissão foi pior do que o de outras crianças e, por isso, não poderia estar significativamente deteriorado na alta hospitalar.

OBJECTIVE: Outcome of patients is determined not only by severity of illness index, but also by the impact of patients' preadmission comorbid status. Therefore, we aimed at evaluating the outcome of patients treated in a pediatric intensive care unit, with special focus on the group of children with chronic diseases. METHODS: Data were obtained prospectively and outcome was assessed according to the Pediatric Overall Performance Category scale for 449 patients in a pediatric intensive care unit of the Split University Hospital. Functional performance was assessed as the preadmission score and the discharge score in patients with neurodevelopmental disabilities, patients with other chronic diseases, and those without chronic disease. RESULTS: The discharge functional status was significantly dependent on the preadmission functional status and on predicted mortality. Children with neurodevelopmental disabilities had the significantly worse baseline score and the significantly smaller deterioration of functional morbidity at discharge compared to children with no chronic disease and children with other chronic diseases. CONCLUSIONS: The Pediatric Overall Performance Category scale has proved its applicability in a small intensive care unit, with a heterogeneous population of patients. It should therefore be considered for regular evaluation of health care quality, as a simple and accurate tool. As opposed to other patients, functional status of children with neurodevelopmental disabilities was markedly influenced by their comorbidity. Their preadmission status was worse than the status of other children, and hence could not significantly deteriorate at discharge.