Identification of intron 1 and intron 22 inversions of factor VIII gene in Serbian patients with hemophilia A

Hemophilia A (HA) is a common X-linked recessive bleeding disease caused by mutations of FVIII gene. Inversion of intron 1 (inv1) and intron 22 (inv22) are recurrent mutations in severe HA, causing 50% of cases. Inv1 has been reported to occur in 2-5% and inv 22 in 45% of severe HA patients. Our objective was to determine, for the first time in Serbia, the frequency of inv1 and inv22 in a group of severe HA patients and to compare these data with those from other countries. Study subjects were 50 HA patients, diagnosed and treated from April 2009 to June 2012 at Mother and Child Health Care Institute of Serbia “Dr Vukan Cupic” (IHS) and Institute for Child and Youth Health Care of Vojvodina (IHV).The presence of inv1 and inv22 was analyzed using Inverse shifting PCR (IS-PCR). Our results revealed that the frequencies of inv1 and inv22 in the cohort of Serbian patients were 6 % and 42% (34% of inv22 type I and 8% of inv22 type II) respectively . These frequencies were in line with those found in other populations. Carrier status analyses of 65 family members (mothers and sisters) showed the de novo inversion of intron 22 in one patient. Genetic Counseling Units of IHS and IHV provide the adequate genetic advice to all HA affected patients and their family members. [Projekat Ministarstva nauke Republike Srbije, br. 173046 i br. 175056

Acute leukemia of childhood: A single institution's experience

The aim of this study was to investigate distribution of immunophenotypic and cytogenetic features of childhood acute leukemia (AL) in the cohort of 239 newly diagnosed patients registered at the leading pediatric oncohematology center in the country during a six-year period (1996-2002). With approximately 60-70% of all childhood AL cases in Serbia and Montenegro being diagnosed and treated in this institution the used data represent a valid research sample to draw conclusions for entire country. On the basis of five phenotypic markers, the distribution of immunological subtypes was as follows: 169 (70.7%) expressed B-cell marker CD19 (137 were CD10 positive and 32 CD10 negative), 37 (15.5%) belonged to T-lineage acute lymphoblastic leukemia (T-ALL) (cyCD3 positive), and 33 (13.8%) were acute myeloblastic leukemia (AML) (CD13 positive and/or CD33 positive in the absence of lymphoid-associated antigens). The ratio of males and females was 1.5:1. Most of the cases were between the ages of 2 and 4, and were predominantly B-lineage acute lymphoblastic leukemia (B-ALL) cases. Another peak of age distribution was observed at the age of 7. The frequency of T-ALL (18% of ALL) was similar to that reported for Mediterranean countries: France (19.4%), Greece (28.1%), Southern Italy (28.3%), and Bulgaria (28.0%). Cytogenetic analyses were performed in 193 patients: 164 ALL and 29 AML. Normal karyotype was found in 57% of ALL and in 55% of AML patients, while cytogenetic abnormalities including structural, numerical, and complex chromosomal rearrangements were found in 43% of ALL and in 45% of AML patients. Our results represent a contribution to epidemiological aspects of childhood leukemia studies

Molecular Analysis of Ring Y Chromosome in a 10-Year-Old Boy with Mixed Gonadal Dysgenesis and Growth Hormone Deficiency

Ring Y chromosome is a very rare chromosomal aberration. The published mixed gonadal dysgenesis (MGD) patients with a ring Y chromosome are short in stature, but are not growth hormone (GH) deficient. We present the molecular cytogenetic and molecular characterization of ring Y chromosome mosaicism in a 10-year-old boy with MGD whose short stature could be explained by the high percentage of cells monosomic for the X-chromosome, but also by the presence of severe GH deficiency. The ring Y chromosome in our patient is a de novo structural aberration. The fathers karyotype was normal

Baseline micronuclei in fetal lymphocytes-a higher frequency in females relative to males

This study provides information concerning base-line micronuclei (MN) and sister-chromatid exchanges (SCEs) in human-fetal lymphocytes. Fetal cord blood (75 samples) were used for purposes of prenatal diagnosis; small aliquots of each were set up into cultures for micronuclei and sister-chromatid exchage analysis. Obtained mean values of base-line MN in fetal lymphocytes are 21.48 +/- 3.52 for males, 24.54 +/- 5.31 for females, respectively. Although small number of fetuses carrying abnormal karyotype was found (6 out of 75), no statistical difference in base-line level of micronuclei between normal versus abnormal karyotype was found. The mean value of SCE is 4.06 +/- 0.77 in female group, 4.03 +/- 0.66 in male group, respectively. As in the case of base-line micronuclei in fetal lymphocytes carrying abnormal karyotype SCEs also are not affected by chromosomal imbalances. There was a lack of correlation between MN and SCE incidence (r = 0.12)

Cytogenetic diepoxybutane sensitivity in Serbian children with Fanconi anemia

Fanconi anemia (FA) is an inherited disorder with aplastic anemia, cancer susceptibility, and hypersensitivity to alkylating agents such as diepoxybutane (DEB). The DEB test is used to screen for FA among patients with bone marrow failure syndromes (BMFS). From February of 2004 to May of 2006, 29 children with BMFS were diagnosed and treated at the Mother and Child Health Care Institute of Serbia (MCHIS). In the examined group, five out of 29 patients (17.2%) were found to have increased DEB-induced chromosome breakage (0.58-2.15 vs. 0.00-0.20 breaks/cell; p<0.001) with no overlap. Our results suggest the importance of this analysis for differential diagnosis and adequate therapy of FA among patients with BMFS

CYTOGENETIC DIEPOXYBUTANE SENSITIVITY IN SERBIAN CHILDREN WITH FANCONI ANEMIA

Abstract – Fanconi anemia (FA) is an inherited disorder with aplastic anemia, cancer susceptibility, and hypersensitivity to alkylating agents such as diepoxybutane (DEB). The DEB test is used to screen for FA among patients with bone marrow failure syndromes (BMFS). From February of 2004 to May of 2006, 29 children with BMFS were diagnosed and treated at the Mother and Child Health Care Institute of Serbia (MCHIS). In the examined group, five out of 29 patients (17.2%) were found to have increased DEB-induced chromosome breakage (0.58-2.15 vs. 0.00-0.20 breaks/cell; p<0.001) with no overlap. Our results suggest the importance of this analysis for differential diagnosis and adequate therapy of FA among patients with BMFS

Chromosomal instability in patients with Fanconi anemia from Serbia

Background/Aim. Fanconi anemia (FA) is a rare hereditary disease in a heterogeneous group of syndromes, so-called chromosome breakage disorders. Specific hypersensitivity of its cells to chemical agents, such as diepoxybutane (DEB), was used as a part of screening among patients with clinical suspicion of FA. The aim of this study was to determine chromosomal instability in patients with FA symptoms in Serbia. Methods. A total of 70 patients with phenotypic symptoms of FA, diagnosed at the Mother and Child Health Care Institute of Serbia “Dr Vukan Čupić”, Belgrade and University Children’s Hospital, Belgrade from February 2004 to September 2011, were included in this study. Cytogenetic instability analysis was performed on untreated and DEBtreated 72 h-cultures of peripheral blood. Results. Ten patients in the group of 70 suspected of FA, showed increased DEB induced chromosome breakage and were classified into the FA group. The range of DEB induced aberrant cells percentages in the FA group was from 32% to 82%. DEB sensitivity of 58 tested patients were bellow FA values (range: 0-6%) (non-FA group), with no overlapping. The remaining two patients showed borderline sensitivity (borderline FA group - FA*), comparing to the healthy controls. Conclusion. This study revealed 10 patients with FA on the basis of cytogenetic analysis of DEB induced chromosome aberrations. Our results are in consistency with those from the literature. Early and precise diagnosis of FA is very important in further treatment of these patients, considering its cancer prone and lethal effects. [Projekat Ministarstva nauke Republike Srbije, br. 173046

Disfunkcija telomera pozitivno koreliše sa incidencom izmena sestrinskih hromatida kod pacijenata obolelih of Fankonijeve anemije

Telomere su nukleoproteinski kompleksi smešteni na krajevima hromozoma koji imaju ključnu funkciju u očuvanju integriteta hromozomskih sekvenci, sprečavajući degradaciju DNK egzonukleazama i neadekvatnu aktivaciju kontrolnih tačaka ćelijskog ciklusa i puteva popravke DNK. TIF metoda (fokusi indukovani telomernom disfunkcijom), predstvalja ko-lokalizaciju faktora DNK repera (53BP1 i γH2AX) i telomera i omogućava kvantifikaciju nefunkcionalnih telomera u različitim tkivima. U primarnim ćelijskim linijama fibroblasta kože devet pacijenata sa sindromom aplazije kostne srži i fenotipom Fankonijeve anemije određivana je bazalna incidenca SCE, T-SCE, γH2AX i TIF. Prosečna učestalost SCE iznoslia je 4.42±0.96 (3.38 do 6.5) po ćeliji, dok je incidenca T-SCE iznoslia 1.91±0.81 (1 do 3.33) po ćeliji. Bazalne vrednosti γH2AX iznosile su 2.27±1.55 (0.8 do 4.2), dok je prosečna učestalosta TIF iznoslia 2.97±1.47 (od 1.44 do 5) po ćeliji. Statistički značajna pozitivna korelacija uočena je između incidence SCE i TIF (0.77 p<0.05), ukazujući da je disfunkcija telomera verovatno nastala mehanizmom homologe rekombinacije.Content available: [https://www.dgsgenetika.org.rs/abstrakti/zbornik-iv-kongres.htm#Rad81

Uticaj antioksidativniih enzima na ukupni radiobiološki odgovor limfocita pacienata obolelih od Fankonijeve anemije

Fankonijeva anemija (FA) je genetičko oboljenje karakterisano progresivnom pancitopenijom i predispozicijom ka malignitetima. Asocirano je sa poremećajima u redoks procesima u ćeliji što ga čini i bolešću oksidativnog stresa. FA ćelije ispoljavaju veliku osetljivost na DNK klastogene agense, dok su podaci o njihovoj osetljivosti na jonizujuće zračenje kontradiktorni. Cilj ove studije je ispitivanje in vitro radioosetljivosti FA homozigota i heterozigota i utvrđivanje uticaja antioksidativnih enzima na ukupan radiobiološki odgovor ćelija. FA pacijenti ispoljavaju značajno sniženje aktivnosti katalaza, sniženu vrednost superoksid dismutaza i povećani bazalni nivo mikronukleusa u odnosu na kontrolu. Radiorezistentni odgovor na jonizujuće zračenje utvrđen je kod većine pacijenata i praćen je značajno većim procentom zračenjem indukovane apoptoze. Utvrđena je negativna korelacija između aktivnosti katalaza i procenta apoptotskih ćelija. Heterozigoti-roditelji imaju visok procenat zračenjem indukovane apoptoze koja je kod nosioca-majki praćena značajnim sniženjem aktivnosti katalaza. Rezultati pokazuju da snižena vrednost katalaza značajno doprinosi ukupnom radiobiološkom odgovoru ćelija.Content available: [https://www.dgsgenetika.org.rs/abstrakti/zbornik-iv-kongres.htm#Rad36