Genetic Profiling of Pediatric Patients with B-Cell Precursor Acute Lymphoblastic Leukemia.

B-cell precursor acute lymphoblastic leukemia (BCP-ALL) is a heterogeneous leukemia subgroup. It has multiple sub-types that are likely to be classified by prognostic factors. Following a systematic literature review, this study analyzed the genes correlated with BCP-ALL prognosis ( IKZF1, PAX5, EBF1, CREBBP, CRLF2, JAK2, ERG, CXCR4, ZAP70, VLA4, NF1, NR3C1, RB1, TSLP, ZNRF1, and FOXO3A) , specifically their nucleotide variations and expression profiles in pediatric BCP-ALL samples. The study included 45 pediatric BCP-ALL patients with no cytogenetic anomaly and a control group of 10 children. The selected genes' hot-spot regions were sequenced using next-generation sequencing, while Polymorphism Phenotyping v2 and Supplemental Nutrition Assistance Program were used to identify pathogenic mutations. The expression analysis was performed using quantitative real-time polymerase chain reaction. The mutation analysis detected 328 variants (28 insertions, 47 indels, 74 nucleotide variants, 75 duplications, and 104 deletions). The most and least frequently mutated genes were IKZF1 and CREBBP , respectively. There were statistically significant differences between patients and controls for mutation distribution in eight genes ( ERG, CRLF2, CREBBP, TSLP, JAK2, ZAP70, FOXO3A, and NR3C1 ). The expression analysis revealed that JAK and ERG were significantly overexpressed in patients compared with controls (respectively, p = 0.004 and p = 0.003). This study combined genes and pathways previously analyzed in pediatric BCP-ALL into one dataset for a comprehensive analysis from the same samples to unravel candidate prognostic biomarkers. Novel mutations were identified in all of the studied genes.

[Investigation of Humoral and Cellular Immunity in TOBB ETÜ Hospital Workers After Three Doses of CoronaVac Vaccination and after One Dose of Comirnaty Vaccination Following Two Doses of CoronaVac]. / TOBB ETÜ Hastanesi Çalisanlarinda Üç Doz CoronaVac ile Iki Doz CoronaVac ve Bir Doz Comirnaty Asisi Sonrasi Humoral ve Hücresel Immünite Düzeylerinin Incelenmesi.

The Coronavirus disease 2019 (COVID-19) pandemic still continues around the world by making peaks with different variants. In the fight against COVID-19, vaccination is currently the only protective measure. In Turkey, vaccination started firstly in healthcare workers on 13 January 2021 with the CoronaVac (Sinovac Biotech, Chinese) vaccine, and booster doses were administered with the CoronaVac and Comirnaty vaccines in July 2021. In this cross-sectional study, it was aimed to determine the humoral and cellular immunity levels of the employees of the TOBB ETU Hospital 2.5-3.5 months after three doses of vaccination and identify the effective factors. With the power analysis that was conducted with the G*Power software, it was determined to be suitable to include 40 TOBB ETU Hospital workers who had their third dose with the CoronaVac vaccine and 60 workers of the same hospital who had their third dose with the Comirnaty (Pfizer Biontech, Germany) vaccine, and age- and sex-matching was considered by selecting 60 workers randomly from among 223 workers who had Comirnaty as their third dose. The study excluded individuals who had a previous COVID-19 infection or had a positive PCR test result. After collecting blood samples on 18-22 October 2021, severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) immunoglobulin G levels were studied with the chemiluminescence immunoassay method using a Beckman-Access device (Beckman Coulter Inc., CA, USA) for humoral immunity, and T-SPOT test (T-SPOT. COVID SARS-CoV-2 test-Oxford Immunotec, Singapur) was carried out for cellular immunity. Additionally, demographic data, habits, personal information such as weight and height, reasons for choosing the vaccine and post-vaccine side effects were obtained through the data collection form. Data collection was completed on 29 November 2021. As a result, among the healthcare workers who had received their third dose of vaccine with Comirnaty and CoronaVac; while 25% of those who received three doses of CoronaVac had humoral immunity, 93.3% humoral immunity was determined in the heterogeneous vaccine group with two doses of CoronaVac and one dose of Comirnaty vaccine (p<0.05). Cellular immunity was found to be higher (88.3%, 70%) in those who received the heterologous vaccine than those who completed three doses of the same vaccine (p<0.05). In the heterogeneous vaccination group consisting of the healthcare workers who had their third vaccine dose with Comirnaty and those who had it with CoronaVac, humoral and cellular immunity levels were found high. Among the participants whose humoral immunity was found negative, cellular immunity was present in 75% of those who had Comirnaty as their third dose and 63.3% of those who had CoronaVac. While no significant relationship was found between immunity levels and age, sex or body mass index (BMI), the levels of both forms of immunity were lower in those who had chronic diseases. Among the participants, physicians preferred the third dose of Comirnaty vaccine at a rate of 71.4% and laboratory workers preferred this vaccine at a rate of 80% according to the workplace. In the third dose vaccine preference reasons, it was stated that 55% would continue with the same vaccine, experience fewer side effects, 44% provide more immunity and 9% do not prevent going abroad. The incidence of side effects after the third dose of vaccine was 53% higher in those who received the Comirnaty vaccine than in those who received CoronaVac (35%). Based on these data, it was concluded that heterologous vaccination should be preferred in vaccination strategies, and knowing cellular immunity levels is important for the decision. There is a need for more comprehensive and follow-up studies on how long humoral and especially cellular immunity lasts.

Identification of novel TUBB1 variants in patients with macrothrombocytopenia.

Background/aim: Macrothrombocytopenia is an autosomal-dominant disorder characterized by increased platelet size and a decreased number of circulating platelets. The membrane skeleton and the link between actin filaments of the skeleton and microtubules, which consist of alpha and beta tubulin [including the tubulin beta-1 chain (TUBB1)] heterodimers, are important for normal platelet morphology, and defects in these systems are associated with macrothrombocytopenia. Materials and methods: In this study, we sequenced the exons of the TUBB1 gene using DNA isolated from the peripheral blood samples of healthy controls (n = 47) and patients with macrothrombocytopenia (n = 37) from Turkey. The TUBB1 expression levels in fractioned blood samples from patients and healthy controls were analyzed by RT-qPCR and Western blot. Microtubule organization of the platelets in the peripheral blood smears of patients, and in mutant TUBB1-transfected HeLa cells, were analyzed by immunofluorescence staining. Results: A new TUBB1 c.803G>T (p.T178T) variant was detected in all of the control and patient samples. Importantly, we found 3 new heterozygous TUBB1 variants predicting amino acid substitutions: G146R (in 1 patient), E123Q (in 1 patient), and T274M (in 4 patients); the latter variant was associated with milder thrombocytopenia in cancer patients treated with paclitaxel. Ectopic expression of TUBB1 T274M/R307H variant in HeLa cells resulted in irregular microtubule organization. Conclusion: Further clinical and functional studies of the newly identified TUBB1 variants may offer important insights into their pathogenicity in macrothrombocytopenia.

Genetic Analysis of Afibrinogenemia and Hypofibrinogenemia: Novel Mutations in the FGB Gene in the Turkish Population.

INTRODUCTION: Congenital afibrinogenemia is a rare autosomal recessive disorder characterized by bleeding that varies from mild to severe and by complete absence or extremely low levels of plasma and platelet fibrinogen. Hypofibrinogenemia is characterized by fibrinogen levels <1.5 g/L. OBJECTIVE: In this study, we analyzed fibrinogen beta chain gene mutations in Turkish afibrinogenemia and hypofibrinogenemia patients. METHODS: We evaluated 20 afibrinogenemia and hypofibrinogenemia patients and 80 healthy controls. We have sequenced all exons of the FGB gene using the DNA isolated from the peripheral blood samples of patients and controls. RESULTS AND CONCLUSION: We found a nonsense mutation in exon 4 at nucleotide 630 that encoded serine amino acid, and in the same exon a missense mutation of T to C at nucleotide 647, resulting in a transition from leucine to proline (p.L198P) in a child with hypofibrinogenemia. These mutations have been shown for the first time in the same patient of Turkish descent. Furthermore, there was a novel heterozygous guanine-to-adenine nucleotide change in exon 3. This caused the change of arginine amino acid to threonine amino acid at position 136 (p.A136T) in a protein, which has not been described in the literature before.

Effects of Ankaferd Hemostat on Helicobacter pylori strains and antibiotic resistance

Background/aim: Ankaferd hemostat (ABS; Ankaferd blood stopper, Istanbul, Turkey) is a folkloric medicinal plant extract. The aim of this study was to determine the effect of Ankaferd hemostat (ABS) on the fate of Helicobacter pylori strains. The study also aims to determine alterations in the antimicrobial resistance of three different H. pylori strains in response to ABS exposure. Materials and methods: H. pylori Strain 1 was obtained from the culture collection ATCC 43504 and passaged three times for viability. Strain 2 was isolated from a gastric ulcer patient and Strain 3 was isolated from a gastritis patient. 1% of ABS was added to all of the strains and antimicrobial susceptibility was observed on 30 and 60 min after application. Results: The efficacy of ABS solutions in achieving significant logarithmic reduction in foodborne pathogens of H. pylori was observed in this study. This study showed that ABS has antibacterial (Anti-H. pylori) effects. Conclusion: Our present study indicated, for the first time, that ABS could act against H. pylori. ABS is clinically used for the management of GI bleeding due to benign and malignant GI lesions. Thus, the possible anti-H. pylori effect of ABS shall expand the therapeutic spectrum of the drug in GI lesions in relation to H. pylori infection such as peptic ulser disease (PUD) and lymphoid tissue (MALT) lymphomagenesis.

Frequency of thiopurine S-methyltransferase gene variations in Turkish children with acute leukemia.

Akin DF, Aslar-Öner D, Kürekçi E, Akar N. Frequency of thiopurine S-methyltransferase gene variations in Turkish children with acute leukemia. Turk J Pediatr 2018; 60: 147-152. In this study we aim to determine the genotype distribution and allele frequencies of common TPMT (*2, *3A, *3B and *3C) polymorphisms in Turkish children with acute leukemia. The study population consisted of 169 patients aged between 1 and 15 years who were admitted to Losante Pediatric Hematology and Children`s Hospital with the diagnosis of acute leukemia. Genotyping of TPMT polymorphisms was screened with real-time PCR using fluorescence melting curve detection analysis. We found that the frequencies of four allelic variants of TPMT are *2 (238 G > C) (0,0%), *3A (460G > A and 719A > G) (1.7%), *3B (460G > A) (1,7%) and *3C (719A > G) (2.4%). Frequency of TPMT alleles increases the efficacy of leukemia treatment. Thus, TPMT genotyping can be useful for optimizing 6-MP therapy.

Frequency of vitamin K oxidoreductase complex subunit-1 (VKORC1) polymorphisms and warfarin dose management in patients with venous thromboembolism.

Warfarin works by inhibiting VKORC1, so polymorphisms of this gene modify the required drug dose. The aim of this study is to examine the relation between therapeutic weekly dose of warfarin and C1173T/G1639A polymorphism of VKORC1 in patients with VTE. Seventy-five patients with VTE were enrolled. Weekly warfarin doses and time (day) to reach therapeutic INR were evaluated retrospectively along with VKORC1-C1173T and G1639A alleles. The mean weekly warfarin dose was lower and time to reach therapeutic INR was shorter in homozygote alleles (AA and TT) (p < 0.05). The multivariate regression model was produced, R2 = 0.05% for age (p = 0.04), R2 = 6% for VKORC1 (p = 0.03), the model for estimating warfarin dose R2 = 17% (p > 0.05). In particular, patients who need overdose of warfarin or whose bleeding score is high, study of these polymorphisms can be considered.

A finely resolved phylogeny of Y chromosome Hg J illuminates the processes of Phoenician and Greek colonizations in the Mediterranean.

In order to improve the phylogeography of the male-specific genetic traces of Greek and Phoenician colonizations on the Northern coasts of the Mediterranean, we performed a geographically structured sampling of seven subclades of haplogroup J in Turkey, Greece and Italy. We resequenced 4.4 Mb of Y-chromosome in 58 subjects, obtaining 1079 high quality variants. We did not find a preferential coalescence of Turkish samples to ancestral nodes, contradicting the simplistic idea of a dispersal and radiation of Hg J as a whole from the Middle East. Upon calibration with an ancient Hg J chromosome, we confirmed that signs of Holocenic Hg J radiations are subtle and date mainly to the Bronze Age. We pinpointed seven variants which could potentially unveil star clusters of sequences, indicative of local expansions. By directly genotyping these variants in Hg J carriers and complementing with published resequenced chromosomes (893 subjects), we provide strong temporal and distributional evidence for markers of the Greek settlement of Magna Graecia (J2a-L397) and Phoenician migrations (rs760148062). Our work generated a minimal but robust list of evolutionarily stable markers to elucidate the demographic dynamics and spatial domains of male-mediated movements across and around the Mediterranean, in the last 6,000 years.

Enlarging the gene-geography of Europe and the Mediterranean area to STR loci of common forensic use: longitudinal and latitudinal frequency gradients.

BACKGROUND: Tetranucleotide Short Tandem Repeats (STRs) for human identification and common use in forensic cases have recently been used to address the population genetics of the North-Eastern Mediterranean area. However, to gain confidence in the inferences made using STRs, this kind of analysis should be challenged with changes in three main aspects of the data, i.e. the sizes of the samples, their distance across space and the genetic background from which they are drawn. AIM: To test the resilience of the gradients previously detected in the North-Eastern Mediterranean to the enlargement of the surveyed area and population set, using revised data. SUBJECTS AND METHODS: STR genotype profiles were obtained from a publicly available database (PopAffilietor databank) and a dataset was assembled including >7000 subjects from the Arabian Peninsula to Scandinavia, genotyped at eight loci. Spatial principal component analysis (sPCA) was applied and the frequency maps of the nine alleles which contributed most strongly to sPC1 were examined in detail. RESULTS: By far the greatest part of diversity was summarised by a single spatial principal component (sPC1), oriented along a SouthEast-to-NorthWest axis. The alleles with the top 5% squared loadings were TH01(9.3), D19S433(14), TH01(6), D19S433(15.2), FGA(20), FGA(24), D3S1358(14), FGA(21) and D2S1338(19). These results confirm a clinal pattern over the whole range for at least four loci (TH01, D19S433, FGA, D3S1358). CONCLUSIONS: Four of the eight STR loci (or even alleles) considered here can reproducibly capture continental arrangements of diversity. This would, in principle, allow for the exploitation of forensic data to clarify important aspects in the formation of local gene pools.

Antibacterial Activities of Ankaferd Hemostat (ABS) on Shiga Toxin-Producing Escherichia coli and Other Pathogens Significant in Foodborne Diseases.

OBJECTIVE: Ankaferd hemostat (Ankaferd Blood Stopper®, ABS)-induced pharmacological modulation of essential erythroid proteins can cause vital erythroid aggregation via acting on fibrinogen gamma. Topical endoscopic ABS application is effective in the controlling of gastrointestinal (GI) system hemorrhages and/or infected GI wounds. Escherichia coli O157:H7, the predominant serotype of enterohemorrhagic E. coli, is a cause of both outbreaks and sporadic cases of hemorrhagic colitis. The aim of this study is to examine the effects of ABS on 6 different Shiga toxigenic E. coli serotypes including O26, O103, O104, O111, O145, and O157 and on other pathogens significant in foodborne diseases, such as Salmonella Typhimurium, Campylobacter jejuni, and Listeria monocytogenes, were also assessed. MATERIALS AND METHODS: All strains were applied with different amounts of ABS and antimicrobial effect was screened. S. Typhimurium groups were screened for survival using the fluorescence in situ hybridization technique. RESULTS: The relative efficacy of ABS solutions to achieve significant logarithmic reduction in foodborne pathogens E. coli O157:H7 and non-O157 serogroups and other emerging foodborne pathogens is demonstrated in this study. ABS has antibacterial effects. CONCLUSION: Our present study indicated for the first time that ABS may act against E. coli O157:H7, which is a cause of thrombotic thrombocytopenic purpura, hemolytic-uremic syndrome, and hemorrhagic colitis. The interrelationships between colitis, infection, and hemostasis within the context of ABS application should be further investigated in future studies.

Spatially Explicit Models to Investigate Geographic Patterns in the Distribution of Forensic STRs: Application to the North-Eastern Mediterranean.

Human forensic STRs used for individual identification have been reported to have little power for inter-population analyses. Several methods have been developed which incorporate information on the spatial distribution of individuals to arrive at a description of the arrangement of diversity. We genotyped at 16 forensic STRs a large population sample obtained from many locations in Italy, Greece and Turkey, i.e. three countries crucial to the understanding of discontinuities at the European/Asian junction and the genetic legacy of ancient migrations, but seldom represented together in previous studies. Using spatial PCA on the full dataset, we detected patterns of population affinities in the area. Additionally, we devised objective criteria to reduce the overall complexity into reduced datasets. Independent spatially explicit methods applied to these latter datasets converged in showing that the extraction of information on long- to medium-range geographical trends and structuring from the overall diversity is possible. All analyses returned the picture of a background clinal variation, with regional discontinuities captured by each of the reduced datasets. Several aspects of our results are confirmed on external STR datasets and replicate those of genome-wide SNP typings. High levels of gene flow were inferred within the main continental areas by coalescent simulations. These results are promising from a microevolutionary perspective, in view of the fast pace at which forensic data are being accumulated for many locales. It is foreseeable that this will allow the exploitation of an invaluable genotypic resource, assembled for other (forensic) purposes, to clarify important aspects in the formation of local gene pools.

Screening of Variations in CD22 Gene in Children with B-Precursor Acute Lymphoblastic Leukemia.

BACKGROUND: CD22 is expressed on the surface of B-cell lineage cells from the early progenitor stage of pro-B cell until terminal differentiation to mature B cells. It plays a role in signal transduction and as a regulator of B-cell receptor signaling in B-cell development. OBJECTIVES: We aimed to screen exons 9-14 of the CD22 gene, which is a mutational hot spot region in B-precursor acute lymphoblastic leukemia (pre-B ALL) patients, to find possible genetic variants that could play role in the pathogenesis of pre-B ALL in Turkish children. METHODS: This study included 109 Turkish children with pre-B ALL who were diagnosed at Losante Hospital for Children with Leukemia. Genomic DNA was extracted from both peripheral blood and bone marrow leukocytes. Gene amplification was performed with PCR, and all samples were screened for the variants by single strand conformation polymorphism. Samples showing band shifts were sequenced on an automated sequencer. RESULTS: In our patient group a total of 9 variants were identified in the CD22 gene by sequencing: a novel variant in intron 10 (T2199G); a missense variant in exon 12; 5 intronic variants between exon 12 and intron 13; a novel intronic variant (C2424T); and a synonymous in exon 13. Thirteen of 109 children (11.9%) carried the T2199G novel intronic variant located in intron 10, and 17 of 109 children (15.6%) carried the C2424T novel intronic variant. CONCLUSION: Novel variants in the CD22 gene in children with pre-B ALL in Turkey that are not present, in the Human Gene Mutation Database or NCBI SNP database, were found.

Qualitative/Chemical Analyses of Ankaferd Hemostat and Its Antioxidant Content in Synthetic Gastric Fluids.

INTRODUCTION: Ankaferd hemostat (ABS) is the first topical haemostatic agent involving the red blood cell-fibrinogen interactions. The antihemorrhagic efficacy of ABS has been tested in controlled clinical trials. The drug induces the formation of an encapsulated complex protein web with vital erythroid aggregation. The aim of this study is to detect the essential toxicity profile and the antioxidant molecules inside ABS. METHODS: The pesticides were analyzed by GC-MS and LC-MS. The determination by ICP-MS after pressure digestion was performed for the heavy metals. HPLC was used for the detection of mycotoxins. Dioxin Response Chemically Activated Luciferase Gene Expression method was used for the dioxin evaluation. TOF-MS and spectra data were evaluated to detect the antioxidants and other molecules. RESULTS: TOF-MS spectra revealed the presence of several antioxidant molecules (including tocotrienols, vitamin E, tryptophan, estriol, galangin, apigenin, oenin, 3,4-divanillyltetrahydrofuran, TBHQ, thymol, BHA, BHT, lycopene, glycyrrhetinic acid, and tomatine), which may have clinical implications in the pharmacobiological actions of ABS. CONCLUSION: The safety of ABS regarding the presence of heavy metals, pesticides, mycotoxins, GMO and dioxins, and PCBs was demonstrated. Thus the present toxicological results indicated the safety of ABS. The antioxidant content of ABS should be investigated in future studies.