Amyloid ß,α-Synuclein and Amyloid ß-α-Synuclein Combination Exert Significant but Different Alterations in Inflammatory Response Profile in Differentiated Human SH-SY5Y Cells.

Neurodegeneration is a condition in which the neuronal structure and functions are altered with reduced neuronal survival and increased neuronal death in the central nervous system (CNS). Amyloid-ß (Aß) is the pathological hallmark of a common neurodegenerative disorder, Alzheimer disease. Parkinson disease and dementia with Lewy bodies are among α-synucleinopathies characterized by abnormal accumulation of insoluble α-synuclein protein. Neuroinflammation is seen in those neurodegenerative disorders regulated by cytokines and chemokines released from neurons, microglia, and astrocytes. Our study aimed to (1) define steady-state levels of cytokines and immune response modulators in SH-SY5Y cells that were differentiated into neuron-like cells and (2) compare the levels of target cytokines in cellular models of neurodegenerative disorders, namely, AD, PD, and DLB-like pathologies. AD, PD, and DLB-like pathologies were established by 6 µM Aß1-42 administration, SNCA (α-synuclein) overexpression, and SNCA overexpression was followed by Aß1-42 treatment, respectively. Alterations in the levels of 40 released inflammatory proteins (IPs) were analyzed by chemiluminescence-based Western/dot blot. Overexpression of human α-synuclein and administration of Aß1-42 significantly changed the profile of IPs secretion, with particularly significant changes in CSF2, CCL5, CXCL8, CXCL10, ICAM1, IL1B, and IL16. Bioinformatics analysis revealed possible interactions between α-synuclein and IL1B. While TGF1, CCL2, TNF, IL10, IL4, and IL1B IPs were associated with Aß 1-42, Aß 1-42 treatment together with α-synuclein, overexpression is associated only with the IL6 protein. Consequently, AD, PD, and DLB-like pathologies might exert significant but different alterations in the inflammatory response.

Palladium complexes containing NNN pincer type ligands and their activities in Suzuki-Miyaura cross coupling reaction.

Five new NNN pincer-type ligands and their palladium complexes were successfully synthesised and characterised by FT-IR, 1H NMR, 13C NMR, and UV-vis analyses. TEM analysis was used to observe the morphological character of the black residues obtained from the fourth cycle of the reusability test. Furthermore, suitable crystals of the N2,N6-bis(2-tert-butylphenyl)pyridine-2,6-dicarboxamide and its palladium complex were elucidated with the X-ray single crystal diffraction method. Both the ligand and its palladium complex crystallise in a monoclinic system with space group P21/c for the H2L4 and C2/c for the palladium complex. The structure of the pincer ligand and its palladium complex were stabilised by intramolecular and intermolecular C-H⋅⋅⋅O, C-H⋅⋅⋅N, and N-H⋅⋅⋅N contacts. A Suzuki-Miyaura cross-coupling reaction between aryl halides and phenylboronic acid was used to assess the catalytic abilities of the palladium pincer complexes. All of the prepared complexes exhibited considerable catalytic activity. However, complexes 4 (Acetonitrile-N2,N6-bis(2-tert-butylphenyl)pyridine-2,6-dicarboxamidopalladium(II)) and 5 (Acetonitrile-N2,N6-bis(2-nitrophenyl)pyridine-2,6-dicarboxamidopalladium(II)) provided almost 100% conversion with nearly 100% yield in the reaction between 4-bromotoluene and phenylboronic acid. Furthermore, these active complexes catalysed the reaction of the sterically hindered and deactivated substrates (1-Bromo-4-izobutylbenzene and 2-bromo-6-methoxynaphthalene) with phenylboronic acid, and complete conversion and yields up to 100% were achieved in a short time with the 2-bromo-6-methoxynaphthalene.

Vitamin D receptor regulates transcription of mitochondrial DNA and directly interacts with mitochondrial DNA and TFAM.

Vitamin D receptor (VDR) is an essential transcription factor (TF) synthesized in different cell types. We hypothesized that VDR might also act as a mitochondrial TF. We conducted the experiments in primary cortical neurons, PC12, HEK293T, SH-SY5Y cell lines, human peripheral blood mononuclear cells (PBMC) and human brain. We showed that vitamin D/VDR affects the expression of mitochondrial DNA (mtDNA) encoded oxidative phosphorylation (OXPHOS) subunits. We observed the co-localization of VDR with mitochondria and the mtDNA with confocal microscopy. mtDNA-chromatin-immunoprecipitation and electrophoretic mobility shift assays indicated that VDR was able to bind to the mtDNA D-loop site in several locations, with a consensus sequence "MMHKCA." We also reported the possible interaction between VDR and mitochondrial transcription factor A (TFAM) and their binding sites located in close proximity in mtDNA. Consequently, our results showed for the first time that VDR was able to bind and regulate mtDNA transcription and interact with TFAM even in the human brain. These results not only revealed a novel function of VDR, but also showed that VDR is indispensable for energy demanded cells.

Association of Maternal Vitamin B12 Status With Infant Findings and Neurodevelopment in Vitamin B12-Deficient Breast-fed Babies.

Few studies have examined the association between maternal vitamin B12 status and their breast-fed infants' findings. The objective of this study was to analyze the association of maternal B12 status with infant findings including neurodevelopmental outcome in breast-fed babies with B12 deficiency. Correlation analyses between the laboratory findings of infants with B12 deficiency (n=120) and their mothers were performed and the association of maternal B12 status with infant findings including the Denver-II developmental screening test (DDST II) results was examined. There was a significant correlation between infant and maternal B12 levels (r=0.222; P=0.030), and between infant and maternal homocysteine (Hcy) levels (r=0.390; P<0.001). Among the babies 4 months of age or older, maternal Hcy levels were significantly correlated with infant mean corpuscular hemoglobin (r=0.404; P=0.001) and infant mean corpuscular volume (r=0.461; P<0.001). Mothers of infants with abnormal DDST II had lower vitamin B12 (196.9±41.2 vs. 247.0±77.0 pg/mL; P=0.018) and higher Hcy levels (17.3±5.0 vs. 10.7±3.1 µmol/L; P<0.001) than mothers of infants with normal DDST II. A lower maternal vitamin B12 status may be related to impaired neurodevelopment in breast-fed infants with vitamin B12 deficiency. Pregnant and lactating women should be offered easy access to healthy nutrition and vitamin B12 supplements.

COVID-19-associated multisystem inflammatory syndrome in children: Experiences of three centres in Turkey.

BACKGROUND: The pathogenesis and clinical manifestations of the multisystem inflammatory syndrome in children (MIS-C) has not yet been fully elucidated and there is no clear consensus on its treatment yet. OBJECTIVES: To evaluate our patients diagnosed with MIS-C and present them to the literature in order to contribute to the better understanding of this new disease, which entered paediatric practice with the SARS-CoV-2 peak. METHODS: In this study, 17 MIS-C cases diagnosed according to the Centers for Disease Control and Prevention criteria were included. RESULTS: Of the patients, 7 (41.2%) had a comorbidity. Gastrointestinal system involvement was the most prominent in the patients (70.6%). Laparotomy was performed in 3 patients due to acute abdomen. Two patients had neurological involvement. Of the patients, 15 (88.2%) received intravenous immunoglobulin and 13 (76.5%) received both intravenous immunoglobulin and methylprednisolone. Two patients received invasive mechanical ventilation and 4 patients received high flow rate nasal cannula oxygen therapy. One of our patients who needed invasive mechanical ventilation and high vasoactive-inotrope support died despite all supportive treatments including plasmapheresis and extracorporeal membrane oxygenation. CONCLUSIONS: MIS-C picture can have a fatal course and may present with severe gastrointestinal and neurological signs. Unnecessary laparotomy should be avoided.

Targeted Next Generation Sequencing and Diagnosis of Congenital Hemolytic Anemias: A Three Years Experience Monocentric Study.

Congenital hemolytic anemias (CHAs) are heterogeneous and rare disorders caused by alterations in structure, membrane transport, metabolism, or red blood cell production. The pathophysiology of these diseases, in particular the rarest, is often poorly understood, and easy-to-apply tools for diagnosis, clinical management, and patient stratification are still lacking. We report the 3-years monocentric experience with a 43 genes targeted Next Generation Sequencing (t-NGS) panel in diagnosis of CHAs; 122 patients from 105 unrelated families were investigated and the results compared with conventional laboratory pathway. Patients were divided in two groups: 1) cases diagnosed with hematologic investigations to be confirmed at molecular level, and 2) patients with unexplained anemia after extensive hematologic investigation. The overall sensitivity of t-NGS was 74 and 35% for families of groups 1 and 2, respectively. Inside this cohort of patients we identified 26 new pathogenic variants confirmed by functional evidence. The implementation of laboratory work-up with t-NGS increased the number of diagnoses in cases with unexplained anemia; cytoskeleton defects are well detected by conventional tools, deserving t-NGS to atypical cases; the diagnosis of Gardos channelopathy, some enzyme deficiencies, familial siterosterolemia, X-linked defects in females and other rare and ultra-rare diseases definitely benefits of t-NGS approaches.

Missed Diagnosis of ß-Thalassemia Trait in Premarital Screening Due to Accompanying HbA2-Yialousa (HBD: c.82G>T).

The diagnosis of ß-thalassemia (ß-thal) trait is usually based on an elevated HbA2 fraction (3.5% to 8%). Co-inheritance of a δ-globin variant along with ß-globin gene defects can interfere with the diagnosis of ß-thal trait by causing normal HbA2 levels. In this report, we present an infant with ß-thal major whose mother's ß-thal trait was missed twice before due to an accompanying δ-globin mutation (HbA2-Yialousa; HBD: c.82G>T), resulting in a borderline HbA2 level. In an individual with microcytosis and hypochromia but an apparently normal HbA2 level, compound heterozygosity for a δ-globin mutation and a ß-thal mutation should be remembered in the differential diagnosis.

Carbonic anhydrase VA deficiency: a very rare case of hyperammonemic encephalopathy.

Objectives Carbonic anhydrase VA (CAVA) deficiency is a rare autosomal recessive inborn error of metabolism that leads to acute metabolic crises, especially in the neonatal or infantile period. It is caused by a deficiency of the enzyme CAVA, which is encoded by the CA5A gene. Case presentation Fifteen patients with homozygous pathogenic CA5A mutations involving 10 different lesions have been reported in the literature up to date. Main clinical and biochemical features of CAVA deficiency include lethargy, hyperammonemic encephalopathy, metabolic acidosis, elevated lactate and hypoglycemia. In most patients reported so far, a single metabolic decompensation attack has been reported, and they have remained stable thereafter with no further crisis. Conclusions We report the 16th case of CAVA deficiency, who was diagnosed by whole-exome sequencing and showed a typical course of the disease with normal development at 18 months.

A novel homozygous nonsense mutation (p.Y78*) in TMPRSS6 gene causing iron-refractory iron deficiency anemia (IRIDA) in two siblings.

Iron-refractory iron deficiency anemia (IRIDA) is an inherited iron metabolism disorder caused by mutations in TMPRSS6 gene encoding matriptase-2, which results in increased hepcidin synthesis. The hallmarks of the disease are hypochromic microcytic anemia, low transferrin saturation, slightly low or normal ferritin levels in contrast to classic iron deficiency anemia (IDA), inadequate response to oral iron, and only a partial response to parenteral iron. We report here a 6-year-old Syrian boy with unexplained microcytic anemia since one year of age. Genetic analysis of the TMPRSS6 gene revealed a novel homozygous nonsense mutation in exon 3 (c.234C > G; p.Y78* or p.Tyr78*). In the presence of hypochromic microcytic anemia accompanied by atypical iron parameters not in accordance with classic IDA, and inadequate response to iron therapy, IRIDA should be remembered in the differential diagnosis.

Factor 8 Gene Mutation Spectrum of 270 Patients with Hemophilia A: Identification of 36 Novel Mutations

Objective: Hemophilia A (HA) is the most severe X-linked inherited bleeding disorder caused by hemizygous mutations in the factor 8 (F8) gene. The aim of this study is to determine the mutation spectrum of the F8 gene in a large HA cohort from Turkey, and then to establish a phenotype-genotype correlation. Materials and Methods: All HA cases (270 patients) analyzed molecularly in the Ege University Pediatric Genetics Molecular Laboratory between March 2017 and March 2018 were included in this study. To identify intron 22 inversion (Inv22), intron 1 inversion (Inv1), small deletion/insertions, and point mutations, molecular analyses of F8 were performed using a sequential application of molecular techniques. Results: The mutation detection success rate was 95.2%. Positive Inv22 was found in 106 patients (39.3%), Inv1 was found in 4 patients (1.5%), and 106 different disease-causing sequence variants were identified in 137 patients (50.6%). In 10 patients (3.7%), amplification failures involving one or more exonic regions, considered to be large intragenic deletions, were identified. Of 106 different F8 mutations, 36 were novel. The relationship between F8 genotype and inhibitor development was considered significant. Conclusion: A high mutation detection rate was achieved via the broad molecular techniques applied in this study, including 36 novel mutations. With regard to mutation types, mutation distribution and their impact on clinical severity and inhibitor development were found to be similar to those previously reported in other hemophilia population studies.

Altered Transcriptional Profile of Mitochondrial DNA-Encoded OXPHOS Subunits, Mitochondria Quality Control Genes, and Intracellular ATP Levels in Blood Samples of Patients with Parkinson's Disease.

Mitochondrial dysfunctions are significant contributors to neurodegeneration. One result or a cause of mitochondrial dysfunction might be the disruption of mtDNA transcription. Limited data indicated an altered expression of mtDNA encoded transcripts in Alzheimer's disease (AD) or Parkinson's disease (PD). The number of mitochondria is high in cells with a high energy demand, such as muscle or nerve cells. AD or PD involves increased risk of cardiomyopathy, suggesting that mitochondrial dysfunction might be systemic. If it is systemic, we should observe it in different cell types. Given that, we wanted to investigate any disruption in the regulation of mtDNA encoded gene expression in addition to PINK1, PARKIN, and ATP levels in peripheral blood samples of PD cases who are affected by a neurodegenerative disorder that is very well known by its mitochondrial aspects. Our results showed for the first time that: 1) age of onset > 50 PD sporadic (PDS) cases: mtDNA transcription and quality control genes were affected; 2) age of onset <50 PDS cases: only mtDNA transcription was affected; and 3) PD cases with familial background: only quality control genes were affected. mtDNA copy number was not a confounder. Intracellular ATP levels of PD case subgroups were significantly higher than those of healthy subjects. We suggest that a systemic dysregulation of transcription of mtDNA or mitochondrial quality control genes might result in the development of a sporadic form of the disease. Additionally, ATP elevation might be an independent compensatory and response mechanism. Hyperactive cells in AD and PD require further investigation.

Inherited coagulation disorders in Turkish children: A retrospective, single-center cohort study.

OBJECTIVE: This study aims to investigate the distribution, clinical characteristics and outcome of inherited coagulation disorders (ICD) in Turkish children. SUBJECTS AND METHODS: Data from all children (age<18 years) with ICD examined in our center were retrospectively reviewed. RESULTS: There were 403 children with ICD (233 males and 170 females) with a median age of four years at diagnosis. The percentages of von Willebrand disease (vWd), hemophilia and rare bleeding disorders (RBD) were 40 %, 34 % and 26 %, type-1, type-2 and type-3 vWd were 63 % 17 % and 20 %, hemophilia A and B were 84 % and 16 %, and severe, moderate and mild hemophilia were 48 %, 30 % and 22 %, respectively. Factor VII and FXI deficiencies were the most prevalent, comprising 56 % and 22 % of all children with RBD, respectively. Parental consanguinity rates were 72 % in type-3 vWd and 61 % in severe RBD. The overall prevalence of gastrointestinal bleedings was 4.5 % (18/403), intracranial bleeding (ICB) was 4.96 % (20/403), mortality from ICB was 30 % (6/20) and the overall mortality rate was 1.49 % (6/403). No life-threatening bleeding was seen during regular prophylaxis. Chronic arthropathy prevalence in severe hemophilia was 8 % with primary prophylaxis and 53 % with demand therap. Inhibitor prevalence was 14 % in hemophilia-A and 5 % in hemophilia-B. CONCLUSIONS: These data show that vWd is the most common ICD, type-3 vWd and RBD are prevalent due to frequent consanguineous marriages and diagnosis of ICD is substantially delayed in Turkish children. Prophylactic replacement therapy prevents occurrence of life-threatening bleedings and reduces the development of hemophilic arthropathy.

Hemolytic Uremic Syndrome in an Infant with Primary Hyperoxaluria Type II: An Unreported Clinical Association.

A 6-month-old boy presented with acute renal failure, thrombocytopenia, and severe non-immune hemolytic anemia. Infection by Shiga-like toxin-producing Escherichia coli and other causes of microangiopathic hemolysis were ruled out, leading to a diagnosis of atypical hemolytic uremic syndrome (aHUS). Neither pathogenic variants in HUS-associated genes nor anti-factor H antibodies were identified. Copy number variation analysis uncovered 4 copies of complement factor H related genes, CFHR1-CFHR4, conceivably leading to higher than normal levels of the corresponding proteins. However, this abnormality was also found in the healthy relatives, neither explaining the disease nor the excessive complement deposition on endothelial cells detected by an ex-vivo test. Whole-exome sequencing revealed a pathogenic homozygous variant in GRHPR encoding the glyoxylate and hydroxypyruvate reductase. Recessive GRHPR mutations cause primary hyperoxaluria type 2 (PH2). The presence of renal calculi in the patient and elevated oxalate levels in the urine were consistent with the genetic diagnosis of PH2. We hypothesize that, in this patient, hyperoxaluria caused by the GRHPR genetic defect triggered endothelial perturbation and complement activation, which was amplified by impaired factor H regulatory activity due to the increased -CFHR1-CFHR4 copy numbers, resulting in aHUS.

An infant with an extremely rare cobalamin disorder: Methionine synthase deficiency and importance of early diagnosis and treatment.

Kasapkara ÇS, Yilmaz-Keskin E, Özbay-Hosnut F, Akçaboy M, Polat E, Olgaç A, Zorlu P. An infant with an extremely rare cobalamin disorder: Methionine synthase deficiency and importance of early diagnosis and treatment. Turk J Pediatr 2019; 61: 282-285. Functional methionine synthase deficiency can be separated into two classes, cobalamin (Cbl) deficiency type E (CblE) and type G (CblG), which are the result of mutations that affect methionine synthase reductase or methionine synthase, respectively. Deficiency of methionine synthase activity may result in megaloblastic anemia without methylmalonic aciduria and neuromuscular abnormality of varying severity. Delayed milestones, ataxia, cerebral atrophy, muscular hypotonia, neonatal seizures, and blindness have been reported as the associated clinical findings. Early diagnosis and treatment are crucial for a more favorable diagnosis of the affected cases. Herein we report a three-month-old boy with CblG disease who presented with failure to thrive, chronic diarrhea, feeding intolerance, oral ulcers, microcephaly and hypotonia, and showed a dramatic response to treatment. In the first few months of life, megaloblastic anemia accompanied by apparent neurological involvement should direct physicians to order examinations like measurement of total homocysteine and methylmalonic acid levels to detect possible forms of inherited Cbl intracellular metabolism disorders.

Association between Maternal and Infantile Markers of Cobalamin Status During the First Month Post-Delivery.

OBJECTIVE: Exclusively breast-fed infants born to vitamin B12 (cobalamin, cbl)-deficient mothers can develop symptoms within a few months following delivery. The authors aimed to assess the relationship between maternal and infantile markers of cbl status. METHODS: In 240 full-term infants (age, 2-30 d) admitted to Samsun Maternity and Child Health Hospital and their mothers, complete blood count testing and serum cbl, folate and plasma total homocysteine (tHcy) measurements were performed. In the mothers, serum ferritin and holotranscobalamin (holoTC) levels were measured additionally. RESULTS: Among the infants, 146 (60.8%) had cbl deficiency (serum cbl <259 pg/mL), whereas 184 (76.7%) mothers had a low cbl level (serum cbl <300 pg/mL). When cbl deficiency was defined as a serum holoTC level < 40 pmol/L, 152 (63.3%) mothers were found as deficient. In addition, 147 (61.3%) infants had an elevated tHcy level (>10 µmol/L), in 35 (23.8%) of these 147 infants tHcy level being markedly elevated (>20 µmol/L). None of the infants had folate deficiency. In the correlational analysis between maternal and infantile markers associated with cbl status, the strongest correlation was observed between maternal holoTC and infantile tHcy (r = -0.49, p < 0.001), followed by the correlation between maternal tHcy and infantile tHcy (r = 0.47, p < 0.001). The weakest correlations were found between maternal cbl and infantile cbl (r = 0.28, p < 0.001), and between maternal cbl and infantile tHcy (r = -0.25, p < 0.001). CONCLUSIONS: Maternal cbl status is an important determinant of infantile cbl status. Both maternal holoTC and tHcy may assist in predicting infantile cbl status. The finding of high prevalence of maternal and infantile cbl deficiency in this study points towards the need for effective strategies to prevent cbl deficiency in women prior to getting pregnant.

Molecular genetic analysis of the F11 gene in 14 Turkish patients with factor XI deficiency: identification of novel and recurrent mutations and their inheritance within families.

BACKGROUND: Factor XI (FXI) deficiency is an autosomal bleeding disease associated with genetic defects in the F11 gene which cause decreased FXI levels or impaired FXI function. An increasing number of mutations has been reported in the FXI mutation database, most of which affect the serine protease domain of the protein. FXI is a heterogeneous disorder associated with a variable bleeding tendency and a variety of causative F11 gene mutations. The molecular basis of FXI deficiency in 14 patients from ten unrelated families in Turkey was analysed to establish genotype-phenotype correlations and inheritance of the mutations in the patients' families. MATERIAL AND METHODS: Fourteen index cases with a diagnosis of FXI deficiency and family members of these patients were enrolled into the study. The patients' F11 genes were amplified by polymerase chain reaction and subjected to direct DNA sequencing analysis. The findings were analysed statistically using bivariate correlations, Pearson's correlation coefficient and the nonparametric Mann-Whitney test. RESULTS: Direct DNA sequencing analysis of the F11 genes revealed that all of the 14 patients had a F11 gene mutation. Eight different mutations were identified in the apple 1, apple 2 or serine protease domains, except one which was a splice site mutation. Six of the mutations were recurrent. Two of the mutations were novel missense mutations, p.Val522Gly and p.Cys581Arg, within the catalytic domain. The p.Trp519Stop mutation was observed in two families whereas all the other mutations were specific to a single family. DISCUSSION: Identification of mutations confirmed the genetic heterogeneity of FXI deficiency. Most of the patients with mutations did not have any bleeding complications, whereas some had severe bleeding symptoms. Genetic screening for F11 gene mutations is important to decrease the mortality and morbidity rate associated with FXI deficiency, which can be life-threatening if bleeding occurs in tissues with high fibrinolytic activity.