The Spectrum of α-Thalassemia Mutations in Syrian Patients.

α-Thalassemia (α-thal) is a globally prevalent genetic disorder of hemoglobin (Hb) structure where the rate of α-globin chain synthesis is reduced or absent due to the presence of α-globin mutation(s). The aim of this study is to define the spectrum of α-globin gene mutations and evaluate their allele frequency in a group of α-thal carriers. A total of 55 individuals with possible α-thal patients were referred from the thalassemia centers in Syria. They have unexplained hypochromia and microcytosis. All patients were genetically tested for 21 common α-globin gene mutations using reverse hybridization kit. Seven different α-globin gene mutations and 13 different genotypes were detected in 55 patients. The two most frequently encountered mutations were -α3.7 deletion (47.1%) and --MED mutation (21.4%). The most commonly observed genotype was -α3.7/αα (40%), followed by --MED/αα genotype (21.8%). We determined the most common α thalassemia mutations in the Syrian patients. α-Thalassemia mutations with deletions were mostly observed in our study.

Secretion of Recombinant Human Annexin V in Fusion with the Super Folder GFP for Labelling Phosphatidylserine-Exposing Membranes.

Annexin V (ANXV), mostly characterized by its ability to interact with biological membranes in a calcium-dependent manner. ANXV interacts mainly with phosphatidylserine (PS), for that fluorescent ANXV widely produced and used as a sensitive and specific probe to mark apoptotic cells or any PS-containing bilayers membranes. Many reports described the prokaryotic expression of recombinant human ANXV. To overcome some of E. coli expression limitations, we aimed in this work to investigate unconventional alternative expression system in mammalian cells for producing secreted human ANXV in fusion with the super folder green fluorescent protein (sfGFP). HEK239T cells were transfected using polyethylenimine (PEI) and pcDNA-sfGFP-ANXV plasmid. Forty-eight hours post transfection, direct fluorescence measurement, immunoblotting and ELISA confirmed the presence of secreted sfGFP-ANXV in cells supernatant. The yield of secreted 6 × His-tagged sfGFP-ANXV after affinity purification was estimated to be around 2 µg per 1 ml of cells supernatant. The secretion system was proper to produce a fully functional sfGFP-ANXV fusion protein in quantities enough to recognize and bind PS-containing surfaces or liposomes. Besides, biological assays such as flow cytometry and fluorescent microscopy confirmed the capacity of the secreted sfGFP-ANXV to detect PS exposure on apoptotic cells. Taken together, we present mammalian expression as a quick, affordable and endotoxin-free system to produce sfGFP-ANXV fusion protein. The secreted sfGFP-ANXV in eukaryotic system is a promising biotechnological tool, it opens up new horizons for additional applications in the detection of PS bearing surfaces and apoptosis in vitro and in vivo assays.

Genotype/Phenotype Correlation of ß-Thalassemia in Syrian Patients: A Cross-Sectional Study.

ß-Thalassemia (ß-thal) is an inherited blood disorder caused by reduced or absent synthesis of ß-globin chains leading to imbalance of globin chain synthesis. ß0-Thalassemia (ß0-thal), refers to the complete absence of ß-globin chain production on the affected allele. ß+-Thalassemia (ß+-thal) refers to alleles with some residual production of ß-globin chain. We studied the correlation of genotype/phenotype of ß-thal disease in Syrian patients. A cross-sectional study was carried out on 260 patients with ß-thal. Genotyping was determined by a DNA sequencing technique. Routine investigations were performed to assess the complete blood count (CBC), serum ferritin, Hb A2 and Hb F levels. We found that the ß0/ß0 genotype was the most common in our patients followed by ß+/ß+ and ß0/ß+. Patients with ß0/ß0 received transfusions at an earlier age and more frequently when compared to those with ß0/ß+ and ß+/ß+ genotypes. Moreover, patients with ß0/ß0 had higher levels of Hb F and lower levels of Hb A2 compared to those with ß0/ß+ and ß+/ß+ genotypes. All patients with ß-thal intermedia (ß-TI) carry the ß+/ß+ genotype, while all patients with ß0/ß0 and ß0/ß+ genotypes presented with transfusion-dependent ß-thal major (ß-TM).

Hb Knossos (HBB: c.82G > T), ß-globin CD 5 (-CT) (HBB: c.17_18delCT) and δ-globin CD 59 (-a) (HBD: c.179delA) mutations in a Syrian patient with ß-thalassemia intermedia.

BACKGROUND: Beta thalassemia (ß-thal) is an inherited hemoglobin disorder characterized by reduced synthesis of the hemoglobin that results in microcytic hypochromic anemia. ß-Thalassemia intermedia (TI) is a clinical term of intermediate gravity between the carrier state and ß-thalassemia major (ß -TM). CASE PRESENTATION: We describe a 12-year-old male proband originating from Al-Quneitra province - southwest Syria. Hematological investigations revealed, pallor and anemia (Hb 9 g/dl). The mean cell volume (MCV) 64 fL; mean cell hemoglobin (MCH) 21.8 pg. Capillary electrophoresis (CE) electropherogram revealed low level of Hb A1 (36.2%), high level of Hb F (62.2%) and low level of Hb A2 (1.6%). The proband requires blood transfusion occasionally. Direct DNA sequencing and Polymerase chain reaction-restriction fragment length polymorphism (PCR/RFLP) for mutations detection were used. The molecular analysis revealed the presence of rare ß+ Hb Knossos codon 27 (G > T) (HBB: c.82G > T) variant associated with ß0 codon 5 [-CT] (HBB: c.17_18delCT) mutation in beta-globin (ß-globin) gene and δ0 codon 59 [-A] (HBD: c.179delA) mutation in delta-globin (δ-globin) gene. The proband tested negative for the common deletional forms of alpha thalassemia (α-thal). Polymorphism of the Xmn-I locus (HBG2: c.-211C > T) revealed that the proband had a homozygous [TT] for Xmn-1 locus. CONCLUSIONS: To our knowledge, this is the first report of beta thalassemia intermedia due to combination of Hb Knossos /codon 5 [-CT] associated with δ0 codon 59 [-A] in Syrian patient. On the other hand, in Syria, ß-thal carriers who have low level of Hb A2 due to decreased δ-chain production, different δ-thal gene mutations must be screened to avoid the failure diagnosis of ß-thal disease.

First Report on the Coinheritance of α-Thalassemia and a Rare ß-Thalassemia Compound Heterozygosity for the IVS-I-I(G>A)/IVS-II-705(T>G) Mutations in a Syrian Family.

We describe a proband originating from Al-Quneitra Province, Syria. His hematology data was as follows: Hb A 24.7%, Hb F 71.1%, Hb A2 4.2%. Molecular analysis, based on DNA sequencing of the ß-globin gene mutation, showed for the first time a compound heterozygous IVS-I-1 (G>A) (HBB: c.92+1G>A)/IVS-II-705 (T>G) (HBB: c.316-146T>G) on the ß-globin gene. A reverse hybridization technique revealed that the proband was also an α-thalassemia (α-thal) -α3.7 (rightward) deletion carrier. Haplotypes analysis for the seven polymorphic restriction sites showed that the compound heterozygous mutations, IVS-I-1/IVS-II-705, were linked to two haplotypes: I [+ - - - - + +] and VI [- + + - - - +], respectively. Our results showed, for the first time, the presence of rare ß-thalassemia (ß-thal) IVS-II-705 (T>G) mutation associated with IVS-I-1 (G>A). Our findings suggest the presence of these mutations resulted from past migrations.

Description of a rare ß-globin gene mutation, IVS-II-848 (C>A) (HBB: c.316-3C>A) in association with IVS-I-1 (G>A) (HBB: c.92 + 1G>A), observed in a Syrian family: a case report.

ß-Thalassemia (ß-thal) is a hereditary and heterogeneous group of disorders caused by mutations on the ß-globin gene that result in the reduced or non production of ß-globin chains. We report a rare ß-globin mutation, IVS-II-848 (C>A) (HBB: c.316-3C>A), which was found in a female Syrian patient. This mutation was associated with the IVS-I-1 (G>A) (HBB: c.92+1G>A) mutation, and the genotype is a compound heterozygote for IVS-I-1(G>A)/IVS-II-848(C>A). This combination was found for the first time in Syria.

HLA-DQ2 and -DQ8 genotype frequency in Syrian celiac disease children: HLA-DQ relative risks evaluation.

BACKGROUND: Celiac disease (CD) is a common autoimmune disease in Syria which manifesting with inflammation of the small intestine and with various extra intestinal symptoms. The disease is associated with human HLA-DQ genes encoding HLA-DQ2 and DQ8 proteins. METHODS: In this study, 49 children patients of CD and 58 healthy control samples were genotyped for HLA-DQ genes using SSP-PCR technique. Relative risks for different genotypes were also evaluated. RESULTS: The DQB1*0201 allele was the most common in the patients (77.6%) followed by DQB1*0302 allele (10.2%). The highest HLA-DQB risk for CD development was found in patients carriers a DQ2.5/DQ8 genotype (1/10), followed by the patients carriers DQ2.5/DQ2.5 (1/12). CONCLUSION: The significant differences in the frequency of HLA-DQ2 and HLA-DQ8 in Syrian patients in compared with controls and relative risks predicted demonstrated the importance role of these alleles in the development of CD in Syrian children patients.

Description of a Rare ß-Globin Gene Mutation: -86 (C>G) (HBB: c.-136C>G) Observed in a Syrian Family.

We present the description of a ß-thalassemia (ß-thal) -86 (C>G) (HBB: c.-136C>G) mutation in a Syrian family from Damascus, As-Suwayda Province, Syria, who was referred to the laboratory for prenatal diagnosis (PND). The mutation was found in the mother in a homozygous state, while it was in the father and in the amniotic fluid sample in a heterozygous state. This mutation is located at -86 within the proximal CACCC box in the promoter of the ß-globin gene and is possibly linked with a phenotype of ß+-thal. Polymerase chain reaction-restriction fragment length polymorphism (PCR/RFLP) analysis indicated that the -86 mutation was linked with haplotype I [+ - - - - + +]. We propose that Lebanon may be the origin of this mutation. To the best of our knowledge, this is the first report describing this mutation in As-Suwayda Province. These findings provide novel information on the region-specificity of this mutation in southwestern Syria.

Haplotype Analysis of Three Common ß-Thalassemia Mutations in Syrian Patients.

ß-Globin haplotypes were used to investigate the origin of three common ß-globin mutations, IVS-I-110 (G>A); HBB: c.93-21G>A, IVS-I-1 (G>A); HBB: c.92 + 1G>A and codon 39 (C>T); HBB: c.118C > T in Syrian patients. Haplotype analysis was done for 49 unrelated patients with ß-thalassemia (ß-thal) and 20 unrelated healthy subjects by polymerase chain reaction (PCR)-based restriction fragment length polymorphism (RFLP) for the ß-globin gene cluster of the following polymorphic restriction sites: HincII 5' to ε, HindIII 5' to Gγ, HindIII 5' to Aγ, HincII in ψß, HincII 3' to ψß, AvaII in ß, and HinfI 3' to ß. The IVS-I-110 mutation was associated with three haplotypes: I [+ - - - - + +] (79.4%), V [+ - - - - + -] (5.9%) and VII [+ - - - - - +] (14.7%), while, the two mutations IVS-I-1 and codon 39 were be linked to a single haplotype V (100.0%) and II [- + + - + + +] (100.0%), respectively. The normal chromosomes (ßA/ßA) were associated with four haplotypes, I (50.0%), II (7.5%), V (32.5%) and VII (10.0%). In the Syrian population, the IVS-I-110 mutation was associated with multi haplotypes, whereas the IVS-I-1 and codon 39 mutations have a single origin. More studies for the other mutations will be very useful for genetic epidemiological studies in Syria.

Induction of G1-phase cell cycle arrest and apoptosis pathway in MDA-MB-231 human breast cancer cells by sulfated polysaccharide extracted from Laurencia papillosa.

BACKGROUND: Marine algae consumption is linked to law cancer incidences in countries that traditionally consume marine products. Hence, Phytochemicals are considered as potential chemo-preventive and chemotherapeutic agents against cancer. We investigated the effects of the algal sulfated polysaccharide extract (ASPE) from the red marine alga L. papillosa on MDA-MB-231 human breast cancer cell line. METHODS: Flow cytometry analysis was performed to study the cell viability, cell cycle arrest and apoptosis. Changes in the expression of certain genes associated with cell cycle regulation was conducted by PCR real time analyses. Further investigations on apoptotic molecules was performed by ROS measurement and protein profiling. RESULTS: ASPE at low doses (10 µg/ml), inhibited cell proliferation, and arrested proliferating MDA-MB-231 cells at G1-phase. However, higher doses (50 µg/ml), triggered apoptosis in those cells. The low dose of ASPE also caused up-regulation of Cip1/p21 and Kip1/p27 and down-regulation of cyclins D1, D2, and E1 transcripts and their related cyclin dependent kinases: Cdk2, Cdk4, and Cdk6. The higher doses of ASPE initiated a dose-dependent apoptotic death in MDA-MB-231 by induction of Bax transcripts, inhibition of Bcl-2 and cleavage of Caspase-3 protein. Over-generation of reactive oxygen species (ROS) were also observed in MDA-MB-231 treated cells. CONCLUSIONS: These findings indicated that ASPE induces G1-phase arrest and apoptosis in MDA-MB-231 cells. ASPE may serve as a potential therapeutic agent for breast cancer.

Frequency of HLA-DRB1 and HLA-DQB1 Alleles and Haplotype Association in Syrian Population.

The study of Human Leukocyte Antigen (HLA) system is very important in health and diseases. As the HLA loci are the most polymorphic in the human genome, it plays a very important role in the immune responses to self and nonself antigens. In the light of the growing importance of typing the HLA alleles in transplantation, autoimmune diseases, cancer, and many other diseases, we studied 225 unrelated healthy Syrian subjects for their HLA class II genotypes in an attempt to reveal the distribution of the HLA (DRB1-DQB1) alleles in the general Syrian population. Our results revealed that the most common alleles for the DRB1 locus were DRB1*11 (26.4%), DRB1*04 (14%), and DRB1*07 (12%). However, the most frequent alleles for the DQB1 locus were DQB1*03 (40.9%) and DQB1*05 (25.1%). The frequent of two-locus haplotypes carry the most frequent alleles at these loci. The most frequently detected class II ''haplotypes'' are DRB1*11-DQB1*03 (8.9%), DRB1*01-DQB1*05 (3.6%), and DRB1*04-DQB1*03 (2.7%). Compared with other populations, our result, deduced from the analysis of genetic distances and the construction of neighbor-joining (NJ) dendrogram, and principal component analysis (PCA) indicates that Syrians are related to Middle Eastern populations. Our data about the Syrian population will aid researchers in studying the relation of HLA class II with different diseases in a Syrian population and will add to the available international literature associated with these loci.

Generation and characterization of nanobodies against rhGH expressed as sfGFP fusion protein.

Growth hormone (GH) deficiencies are diagnosed in most children with short stature and treated with a long course of administrating expensive and daily doses of recombinant human GH (rhGH or Somatropin®). This work describes for the first time the production of several GH specific nanobodies with great potential in the field of GH production and detection. Nanobodies are the smallest intact antigen binders derived from heavy chain-only antibodies (HCAbs) of camelids. They are very stable, highly soluble and are produced as recombinant proteins in Escherichiacoli at an affordable cost for various biotechnological applications. To increase its solubility and immunogenicity, GH was produced as fusion with superfolder green fluorescent protein (sfGFP) and was used in this form to successfully immunize an adult camel. The active involvement of HCAbs in the specific camel immune response encouraged the preparation of large nanobody "immune" library. Phage display biopanning of this library against GH resulted in the isolation of five interesting and different nanobodies, referred to as NbGH01, 02, 03, 04 and 06. All nanobodies were able to recognize GH in its fusion and free formats and the detection sensitivity ranged from 0.5 to 10 ng/ml in sandwich ELISA. Pure rhGH was successfully purified by affinity chromatography, using immobilized NbGH06, from the cleavage reaction of fusion proteins with the tobaccos etch virus (TEV) protease. These specific molecular binders, especially NbGH06, provide valuable tools for rhGH diagnostic as well as for production purposes.

Prenatal molecular diagnosis of ß-thalassemia and sickle cell anemia in the Syrian population.

Our objective was to evaluate the prenatal diagnosis (PND) of ß-thalassemia (ß-thal) and sickle cell anemia in Syria. Mutations detected from blood of at-risk couples and 55 amniotic fluid samples collected at the second trimester of pregnancy (14-22 weeks' gestation) were characterized. Molecular screening and direct DNA sequencing of the HBB gene was carried out. DNA analyses showed 14 affected fetuses (25.45%), 32 (58.18%) carriers and eight (14.54%) normal fetuses. It appears that 20.0% of individuals carried the sickle cell anemia mutation and 80.0% carried the ß-thal mutation. Thirteen different known mutations were detected in the fetuses. The most common mutations were: IVS-II-1 (G > A), codon 39 (C > T)], IVS-I-110 (G > A), IVS-I-1 (G > A) and IVS-I-5 (G > C). The Hb S [ß6(A3)Glu → Val; HBB: c.20A > T] mutation was the only abnormal hemoglobin (Hb) that was found. The results point to a successful future for PND of ß-thal and sickle cell anemia in Syria, using a rapid and accurate molecular method. We hope that this method will be used as a common application approach to decrease the incidence of ß-thal major (ß-TM).

Molecular update of ß-thalassemia mutations in the Syrian population: identification of rare ß-thalassemia mutations.

ß-Thalassemia (ß-thal) is an autosomal recessive disorder characterized by variable degrees of anemia, bone marrow hyperplasia, splenomegaly, and complications related to the severity of the anemic state. The ß-thalassemias result from mutations in and around the ß-globin gene (HBB) located as a cluster on the short arm of chromosome 11. In Syria, ß-thal is highly prevalent. The main aim of this study was to identify the frequency of HBB mutations in 189 Syrian ß-thal patients and carriers of ß-thal. Out of the 189 patients and carriers recruited in this study, 181 patients had at least one HBB mutation and eight patients did not show any mutation. The 10 most frequent ones constituted 77.5% of all HBB mutations. These mutations in order of frequency were: IVS-I-110 (G > A) (17.0%), IVS-I-1 (G > A) (14.7%), codon 39 (C > T) (14.4%), IVS-II-1 (G > A) (9.8%), codon 8 (-AA) (6.2%), IVS-I-6 (T > C) (5.2%), IVS-I-5 (G > C) (4.9%), codon 5 (-C) (3.2%), IVS-I-5 (G > A) (3.2%) and codon 37 (G > A) (2.2%). Another 21 mutations were less frequent or sporadic. These results provide important tools for adapting a prenatal molecular diagnostic test for the Syrian population.

Compound heterozygosity for Hb C/Hb S (HBB: c.19G>A/HBB: c.20A>T) diseases observed in a Syrian family: a case report.

Hemoglobin S and Hemoglobin C disease is a type of sickle cell disease caused by two mutations at codon 6 of ß-globin gene. These mutations cause changes in the shape of the red blood cells. Little is known about its presence in our region. Case Presentation: The authors describe a case of a Syrian family consisting of father, mother, two daughters, and son. The mother presented with anemia, episodes of fatigue, and extreme pain (vaso-occlusive crisis). Beta and alpha-globin gene mutations have been analyzed using molecular detection methods. The results revealed that, the mother, second daughter, and son were all double heterozygous for hemoglobin C and S associated with the -α3.7 deletion mutation. The husband and the first daughter were found to have the hemoglobin C trait. Discussion: Hemoglobin (Hb) SC has been known to have a higher frequency in black populations and is restricted to persons of West African descent. In our case, all family members had dark brown skin color, and they were all diagnosed with Hb C or Hb SC. The mother, second daughter, and son had the clinical manifestations associated with Hb SC disease, and their values of mean cell volume and mean cell hemoglobin were low due to the presence of the -α3.7 deletion mutation. The husband and the first daughter do not have any serious health problems. Conclusions: To the best of the knowledge, this is the first case of compound heterozygous for hemoglobin C and S to be reported from a Syrian family.

"In House" assays for the quantification of Annexin V and its autoantibodies in patients with recurrent pregnancy loss and in vitro fertilisation failures.

Several studies have been shown that Annexin V (ANXV) autoantibodies concentrations are associated with both early recurrent pregnancy losses (RPLs) or in vitro fertilization failure (IVFf). We investigated the association between ANXV autoantibodies and ANVX levels in RPL, IVFf and normal group women. The study was conducted on 22 female patients with RPLs, 66 patients with IVFf, and 16 normal samples from women who had given birth. ANXV autoantibodies were measured using an ELISA test developed by fixing a homemade recombinant ANXV protein and examined with labeled human antibodies, while ANXV concentrations were measured by a competitive ELISA using a homemade anti ANXV polyclonal antibody. The results showed a clear relationship between the high levels of ANXV autoantibodies and the recurrent abortion. On the other hand, ANXV measurement in those patients showed decreased concentrations compared to normal samples. Negative correlation between ANXV and its autoantibodies levels was reported in almost all patients' samples. Our data supports the possibility that ANXV autoantibodies are a risk factor for reproductive failures associated with both RPLs and/or IVFf and the significant role for ANXV in the maintenance of pregnancy.

Investigation of role of CpG methylation in some epithelial mesenchymal transition gene in a chemoresistant ovarian cancer cell line.

Ovarian cancer is one of the lethal gynecologic cancers. Chemoresistance is an essential reason for treatment failure and high mortality. Emerging evidence connects epithelial-mesenchymal transition (EMT) like changes and acquisition of chemoresistance in cancers. Including EMT, DNA methylation influences cellular processes. Here, EMT-like changes were investigated in cisplatin-resistant A2780 ovarian cancer cells (A2780cis), wherein role of DNA methylation in some EMT genes regulations was studied. Cell viability assay was carried out to test the sensitivity of A2780, and A2780cis human cancer cell lines to cisplatin. Differential mRNA expression of EMT markers using qPCR was conducted to investigate EMT like changes. CpG methylation role in gene expression regulation was investigated by 5-azacytidine (5-aza) treatment. DNA methylation changes in EMT genes were identified using Methylscreen assay between A2780 and A2780cis cells. In order to evaluate if DNA methylation changes are causally underlying EMT, treatment with 5-aza followed by Cisplatin was done on A2780cis cells. Accordingly, morphological changes were studied under the microscope, whereas EMT marker's gene expression changes were investigated using qPCR. In this respect, A2780cis cell line has maintained its cisplatin tolerance ability and exhibits phenotypic changes congruent with EMT. Methylscreen assay and qPCR study have revealed DNA hypermethylation in promoters of epithelial adhesion molecules CDH1 and EPCAM in A2780cis compared to the cisplatin-sensitive parental cells. These changes were concomitant with gene expression down-regulation. DNA hypomethylation associated with transcription up-regulation of the mesenchymal marker TWIST2 was observed in the resistant cells. Azacytidine treatment confirmed DNA methylation role in regulating gene expression of CDH1, EPCAM and TWIST2 genes. A2780cis cell line undergoes EMT like changes, and EMT genes are regulated by DNA methylation. To that end, a better understanding of the molecular alterations that correlate with chemoresistance may lead to therapeutic benefits such as chemosensitivity restoration.

A rare gene variation cap +1 (A>C) (HBB: c. -50A>C) associated with codon 5 (-CT) (HBB: c.17_18delCT) mutation in Syrian family.

BACKGROUND: CAP+1 [A>C] (HBB:c.-50A>C) is a rare silent ß-thalassemia (ß-thal) mutation. Carrier individuals of this mutation show borderline hemoglobin (Hb), mean corpuscular volume (MCV) and Hb A2 levels. This mutation was previously reported in combination with different ß-thalassemia mutations, leading to variable phenotypes. CASE PRESENTATION: Here, we describe for the first time the combination of silent CAP+1 [A>C] (HBB:c.-50A>C) mutation with ß0 codon 5 [-CT] (HBB:c.17_18delCT) mutation in a Syrian proband, leading to beta thalassemia intermedia (TI). CONCLUSIONS: The compound heterozygotes of the silent CAP+1 (A>C) together with another severe beta gene mutation, are phenotypically severe enough to present at an early age and require appropriate therapeutic modalities.

Molecular analysis of the AGXT gene in Syrian patients suspected with primary hyperoxaluria type 1.

BACKGROUND: Characterization of the molecular basis of primary hyperoxaluria type 1 (PH-1) in Syria has been accomplished through the analysis of 90 unrelated chromosomes from 45 Syrians patients with PH-1 from different regions. METHODS: Alanine glyoxylate aminotransferase (AGXT) gene mutations have been analyzed by using molecular detection methods based on the direct DNA sequencing for all exons of the AGXT gene. RESULTS: Seventeen pathogenic mutations were detected in our patients. Six mutations were novels. The three most frequent mutations were c.33_34insC (p.Lys12fs) in Exon 1, c.584 T < G; p.Met195Arg in exon 5 and c.1007 T > A (p.Val336Asp) in exon 10, with a frequency of 33.3%, 12.2%, and 11.1%, respectively. CONCLUSION: DNA sequencing used in this study can offer a useful method to investigate the mutations in Syrian PH-1 patients, and could offer an accurate tool for prenatal diagnosis and genetic counseling.

Effects of ionizing radiation on the viability and proliferative behavior of the human glioblastoma T98G cell line.

OBJECTIVE: Radiotherapy is the traditional therapy for glioma patients. Glioma has poor response to ionizing radiation (IR). Studying radiation-induced cell death can help in understanding the cellular mechanisms underlying its radioresistance. T98G cell line was irradiated with Co60 source by 2 or 10 Gy. MTT assay was used to calculate the surviving fraction. Cell viability, cell cycle distribution and apoptosis assays were conducted by flow cytometry for irradiated and control cells for the 10 Gy dose. RESULTS: The SF2 value for irradiated cells was 0.8. Cell viability was decreased from 93.29 to 73.61%, while, the Sub G0/G1 phase fraction was significantly increased at 10 Gy after 48 h. On the other hand, there was an increase in the percentage of apoptotic cells which reached 40.16% after 72 h at the same dose, while, it did not exceeds 2% for non-irradiated cells. Our results showed that, the T98G cells is radioresistant to IR up to 10 Gy. Effects of irradiation on the viability of T98G cells were relatively mild, since entering apoptosis was delayed for about 3 days after irradiation.