alihossein saberi

In this study, we describe a new missense variant on the β-globin gene in a heterozygous form in a female individual. Standard methods were used to determine red blood cell indices and perform hemoglobin analyses. Molecular studies were performed on the genomic DNA isolated from peripheral blood cells. Beta-globin genes were amplified and sequenced. We report a novel mutation on the β-globin gene (HBB), c.134 C>T; p.S44F variant, in the heterozygote state which was detected in a female of Persian ethnic origin in the Khuzestan province, southern Iran, that we named Hb Narges Lab (HbNL) variant. This mutation was predicted to be disease-causing in all except one in silico prediction tools. This variant was reported for the first time worldwide, had no shown hematological abnormalities but should be considered when inherited in the compound heterozygous form with β- thalassemia (β0-thal) carrier, which might result in the phenotype of thalassemia intermedia.

Hyperlipidemia is one of the main risk factors for coronary artery disease and is defined as abnormal elevation of lipids or lipoproteins in the blood. Pcsk9 is the ninth member of the proprotein convertase family that binds to the LDLR on the surface of the hepatocyte, leading to degradation of LDLR in lysosomes which could cause hyperlipidemia. The present study aimed to analyze the methylation status of pcsk9 promoter in patients with hyperlipidemia.

This case-control study was conducted in 50 patients with hyperlipidemia and 50 healthy controls. DNA isolation from whole blood was performed using salting out procedure. Promoter methylation of the Pcsk9 gene was analyzed by methylation-specific PCR (MSP). Ten MSP products were sequenced to confirm the data obtained.

According to MSP results, methylation pattern of pcsk9 gene promoter displayed an unmethylated status among the patients and control individuals. In other words, no methylation was seen in case and control samples.

The current study showed no significant association between PCSK9 methylation pattern and blood lipid profile level in case group and control group.

Plasma levels of cell-free DNA (cfDNA) are elevated in various clinical conditions, including cancer, myocardial infarction, autoimmune disease, and pregnancy-associated complications. The aim of this study was to determine the level of cfDNA to assess the potential of cfDNA as a biomarker for screening and diagnosis and also its effectiveness in the treatment of patients with acute lymphoblastic leukemia (ALL).

Overall, 40 individuals (20 healthy volunteers and 20 patients with ALL) were examined. For quantitative analysis of cfDNA, plasma purified DNA was subjected to real-time PCR amplification of the beta-globin gene. Quantification of cfDNA levels was performed at the time of diagnosis and after treatment with a common protocol.

At diagnosis, in all samples, plasma levels of cfDNA were significantly elevated compared to those of healthy controls after treatment. The high levels of cfDNA decreased and returned to normal after treatment.

Data showed that despite the significant cfDNA concentration increment in patients than the control group, its detection and treatment potential should be more studied: however, it still can be a useful marker for screening of diseases, such as hematological neoplasms.

Various blood diseases are caused by mutations in the FANCA, FANCC, and ITGA2B genes. Exome sequencing is a suitable method for identifying single‑gene disease and genetic heterogeneity complaints.

Among families who were referred to Narges Genetic and PND Laboratory in 2015‑2017, five families with a history of blood diseases were analyzed using the whole exome sequencing (WES) method.

We detected two novel mutations (c.190‑2A>G and c.2840C>G) in the FANCA gene, c. 1429dupA mutation in the FANCC gene, and c.1392A>G mutation in the ITGA2B gene. The prediction of variant pathogenicity has been done using bioinformatics tools such as Mutation taster PhD‑SNP and polyphen2 and were confirmed by Sanger sequencing.

WES could be as a precise tool for identifying the pathologic variants in affected patient and heterozygous carriers among families. This highly successful technique will remain at the forefront of platelet and blood genomic research.

Hemoglobin (Hb) Alesha is a rare and very unstable Hb variant, resulting in disruption of the heme pocket and producing severe hemolysis in heterozygous statues. In this study, we describe the first report of this variant in an Iranian boy originated from south of Iran with severe hemolytic anemia and mild splenomegaly.

A six-year-old boy from Khuzestan Province and his parents were studied. Gap-PCR and direct sequencing were performed to detect the a-globin gene deletions and β-globin gene mutations, respectively.

The subject had a sporadic mutation GTG to ATG (Val [valine]>Met [methionine]) at codon 67 in heterozygous form on β-globin gene, which was not detected in his parents.

Since both parents proved to be normal, this Hb variant could be considered as a de novo mutation, which is highly useful for prenatal diagnosis.

  Niemann-Pick diseases (NPD) is an autosomal recessive inherited lysosomal lipid storage disorder which occurs due to a defect in cellular cholesterol trafficking, leading to excess lipid accumulation in multiple organ systems such as the brain, lungs, spleen and liver. SPMD1-associated disease includes classic infantile and visceral NPD type A and B respectively. Type C NPD is subacute or juvenile. Sanger sequencing of the candidate genes for NPD were performed followed by bioinformatic analysis to confirm the types of NPD and to identify novel mutations. All patients underwent full clinical assessment. In this case series, we present two cases with NPD type A, six cases with NPD type B, and 11 cases with type C with various enzymatic defects identified in these cases. Within these 19 patients we present seven previously reported mutations and 10 novel mutations causing NPD. Our report demonstrates that NPD has a variable age of onset and can present early in life. In this study, we investigated the clinical and genetic manifestations of a large Iranian cohort. Understanding the variable presentation of NPD will allow for clinicians to have a high index of suspicion for the disease.

In this study, a high-performance liquid chromatography method was developed to determine the concentration of gliclazide in plasma. The Photodiode array (PDA) detector allows for the determination of purity.
Subjects and The mobile phase used in this study was phosphate buffer with pH = 3.5 and acetonitrile (HPLC grade) at a ratio of 1: 1 was run isocraticaly through a C18 analytical column. The UV detection was done at 253 nm. Analytical run time was less than 10 min.
In the optimum conditions, the calibration curve in the area of ​50 to 400 ng/ml, linear with the equation y = 28.029x 212.74 and 0.998 = R2. Gliclazide was detected within 3.6 minutes by photodiode detector. The limit of detection (LOD) and the limit of quantitative (LOQ) in this method are calculated as 23 ng/ml and 71 ng/ml, respectively.
The evaluation of the accuracy and precision of method showed that this method has the ability to determine the amount of gliclazide in plasma, and the method is reliable and fast.

The increase of HbF in some beta-thalassemia and sickle cell patients ameliorates the disease severity. Therefore, understanding the reasons behind the increasing of fetal hemoglobin is needful. The molecular explanation of fetal hemoglobin increment might be correlated with the genes which play a regulatory role in expression of beta-globin cluster genes. In this study, the likelihood of polymorphisms of BCL11A and HBS1L-MYB regions to cause HbFelevation was studied.
Subjects and In this investigation, 50 thalassemia patients with high level of fetal hemoglobin and 50 healthy individuals as control group were selected from Khuzestan. The allele frequency of rs28384513, rs766432, rs11886868 and rs4895441 polymorphisms were assessed by using PCR-RFLP and PCR- sequencing methods.
The frequency of C allele in rs766432 (A/C), C allele in rs11886868 (C/T) and A allele in rs28384513 (A/C) were significantly high among the patients with increased level of HbF. Whereas, the frequency of rs4895441 (A/G) polymorphism had no significant differences compared with the patients with high level of HbF and control group.
Based on this study, rs766432 (A/C), rs11886868 (C/T) and rs28384513 (A/C) polymorphisms are correlated with HbF increased and they can be used to predict the clinical features of fetus in case of bearing the major or intermediate forms of thalassemia. However, to confirm these results these markers need to be studied by applying a larger number of samples.

β-thalassemia is one of the most widespread disease in the world, including Iran. In this study, we reported, for the first time, A 290-bp β-globin gene deletion in the south of Iran.
Four individuals from three unrelated families with Arabic ethnic background were studied in Khuzestan Province. Red blood cell indices and hemoglobin analysis were carried out according to the standard methods. Genomic DNA was obtained from peripheral blood cells by salting out procedures. β-globin gene amplification, multiplex ligation-dependent probe amplification (MLPA) and DNA sequencing were performed.
The PCR followed by sequencing and MLPA test of the β-globin gene confirmed the presence of a 290-bp deletion in the heterozygous form, along with -88C>A mutation. All the individuals had elevated hemoglobin A2 and normal fetal hemoglobin levels.
This mutation causes β0-thalassemia and can be highly useful for prenatal diagnosis in compound heterozygous condition with different β-globin gene mutations.

The recent investigations have rendered microRNAs (miRs) as a novel biomarker in cancer research. In fact, alteration in miR expression may be associated with tumor suppression, tumorigenesis, metastasis, and poor prognosis in human breast cancer (BC).
The aim of this clinical experimental study was to measure the miR-328 expression level in breast cancer tissues, at first. Then, we tried to find out any possible correlation between miR-328 and prognostic and predictive biomarkers in BC. Both of these two objectives were investigated for the first time; and we did not find any similar survey measuring the expression level of miR-328 in both tumor and non-tumor breast tissues. This research was conducted in Iran (Ahvaz, Khuzestan), between December 2013 and April 2014. Furthermore, we did not find any previous document investigating the correlation between miR-328 expression level and prognostic factors in BC. Due to the lack of similar studies intending to measure the expression level of miR-328 in tumor and adjacent non-tumor tissues, we decided to carry out a pilot study.
We measured the expression level of miR-328 by Poly (A) real-time PCR based on SYBR Green-I in 28 fresh samples of BC tissues and 28 samples of normal adjacent tissues, including invasive ductal carcinoma (IDC), invasive lobular carcinoma (ILC), and ductal carcinoma in situ (DCIS). We tried to attribute the results to clinicopathologic features such as status of estrogen and progesterone receptors (ER/PR), HER2/neu (HER2), P53 and also Ki67 labeling (Ki67-LI).
The results showed that the miR-328 median level of expression was 0.88 (2-ΔΔCt) (25th-75th percentile, 0.07 - 2.34). It means that the expression level increased in tumor tissues compared to normal adjacent tissues (NATs). However, a statistically significant correlation between the miR-328 median expression level and prognostic factors, including pathologic diagnosis, age, and also the status of ER, PR, HER2, and Ki67-LI was not observed (P > 0.05).
Therefore, it might be possible to consider miR-328 as an oncogene; but not necessarily an oncomiR, in human BC.

Rapid dose assessment using biological dosimetry methods is essential to increase the chance of survival of exposed individuals in radiation accidents.
We compared the expression levels of the FDXR and RAD51 genes at 6 and 18 MV beam energies in human peripheral blood lymphocytes. The results of our study can be used to analyze radiation energy in biological dosimetry.
For this in vitro experimental study, from 36 students in the medical physics and virology departments, seven voluntary, healthy, non-smoking male blood donors of Khuzestan ethnicity with no history of exposure to ionization radiation were selected using simple randomized sampling. Sixty-three peripheral blood samples were collected from the seven healthy donors. Human peripheral blood was then exposed to doses of 0, 0.2, 0.5, 2, and 4 Gy with 6 and 18 MV beam energies in a Linac Varian 2100C/D (Varian, USA) at Golestan hospital in Ahvaz, Iran. After RNA extraction and cDNA synthesis, the expression levels of FDXR and RAD51 were determined 24 hours post-irradiation using the gel-purified reverse transcription polymerase chain reaction (RT-PCR) technique and TaqMan strategy (by real-time PCR).
The expression level of FDXR gene was significantly increased at doses of 2 Gy and 4 Gy in the 6 - 18 MV energy range (P The data suggest that the expression analysis of the FDXR gene, contrary to that of the RAD51 gene, may be suitable for assessment of high-energy X-ray. In addition, RAD51 is not a suitable gene for dose assessment in biological dosimetry.

Maple syrup urine disease (MSUD) is a rare metabolic disorder caused by deficiency in branched chain alpha-keto acid dehydrogenase complex (BCKD).
In this study, the coding regions and flanking splice sites of the BCKDHA, BCKDHB, DBT and DLD genes have been sequenced in an Iranian 3 years old girl.
A novel homozygous mutation (p.Glu330Lys) was detected in the BCKDHB gene. In silico analysis showed significant change in the 3-D protein Structure.
This alteration probably affects the structure and function of the E1β subunit of BCKD complex.

A sub-population of tumor cells termed cancer stem cells (CSCs) has an important role in tumor initiation, progression, and recurrence. Selecting a suitable procedure for isolation and enrichment of CSCs is the biggest challenge in the study of CSCs. In the present study, the role of the combination of stem cell culture medium and collagen type-I was evaluated for successful isolation and enrichment of HT-29 CSCs.

HT-29 cells were cultured in serum-containing medium (parental culture medium: Medium + 10% fetal bovine serum) and serum-free medium (stem cell culture medium); both on collagen-coated plates. Spheres forming ability and CD133 expression, as a potential marker of colorectal CSCs, were evaluated in two culture mediums.

The results show spheroids usually give rise completely within 15 days in the stem cell culture medium on the collagen-coated plate. CD133 expression in spheroid cells (84%) is extensively higher than in parental cells (25%). Moreover, relative to parental cells, spheroid cells were more radioresistance.

Finding of this study suggested that CSCs derived from colon cancer cell line (HT-29) can be propagated and form colonospheres in serum-free culture medium on collagen type-I. According to maintenance of their original phenotype in these conditions, it seems serum-free culture medium on collagen type-I is a suitable way to drug screening of HT-29 CSCs.

Mean inactivation dose is a useful radiobiological parameter for the comparison of human cell survival curves. Given the importance and accuracy of these parameters, in the present study, the radio sensitivity enhancement of colon cancer (HT-29) cells in the presence of gold nanoparticles (GNPs) were studied using the mean inactivation dose (MID). Naked-GNPs with 50 nm diameters were incubated with HT-29 cells. The cytotoxicity and uptake of these particles on HT-29 cells were assessed. After determining the optimum GNPs concentration, the cells were incubated with gold nanoparticle for 24 hours. The change in the MID value as well as the radio sensitization enhancement under irradiation with 9 MV X-ray beams in the presence of GNPs were evaluated by multiple (3-(4, 5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium)MTS assay. Cell survival in the presence of GNPs was more than 90% and the maximum uptake of GNPs was observed at 60 µM of gold nanoparticles. In contrast, in the presence of GNPs combined with radiation, cell survival and MID value significantly decreased, so that the radio sensitization enhancement was 1.4. Due to the significant reduction in the mean inactivation dose of colon cancer cells in the presence of gold nanoparticles, it seems that GNPs are suitable options to achieve a new approach in order to improve radiotherapy efficiency without increasing the prescribed radiation dose.

Autosomal dominant polycystic kidney disease (ADPKD) is one of the most common genetic kidney disorders. Genetic studies have demonstrated an important allelic variability among patients but very few data are known about the genetic variation in Iranian populations. In this study, in order to verify the ADPKD in a patient with some clinical symptoms and study the variations of the PKD1 gene for the first time in Iranian population, the PKD1 gene was entirely sequenced. Coding exons analysis of PKD1 by exon direct sequencing was performed. Molecular genetic testing found a novel mutation in the patient. It was a missense mutation CAT > GAT at position 3311 in exon 30 of PKD1. CAT > GAT causes the conversion of amino acids histidine to argenine and change the transmembrane domain and proper function of the polycystin 1 protein.

Wolf-Hirschhorn syndrome (WHS) is a disorder that affects many parts of the body. The major features of this condition include specific craniofacial malformations, delayed growth and development, intellectual disability and seizures. Here, we report a case of WHS: a 27-month-old girl with a microdeletion at distal part of short arm of chromosome 4. She had striking clinical features of WHS and had an apparently normal karyotype. Array comparative genomic hybridization performed on the DNA extracted from peripheral blood revealed loss of 1.7Mb at 4q16.3-q15.3. Taken together, this data suggests that a patient with strong clinical suspicion of chromosome abnormality and normal conventional karyotype analysis should be further evaluated by molecular cytogenetic techniques such as array comparative genomic hybridization (aCGH) or fluorescence in situ hybridization (FISH).

In this study, a new alpha globin gene mutation on the α2-globin gene is reported. This mutation resulted in a Lys > stop codon substitution at position 127 which was detected in four individuals (three males and one female). DNA sequencing revealed this mutation in unrelated persons in Khuzestan province, Southwestern Iran of Lor ethnicity. This mutation caused no severe hematological abnormalities in the carriers. From the nature of substituted residues in α2-globin, it is widely expected that this mutation leads to unstable and truncated protein and should be detected in couples at risk for α-thalassemia.

Renal tubular acidosis (RTA) is a rare disorder. It has four clinical types, which ‘Type 3 RTA’ demonstrating a mixed pattern of tubular dysfunction. The causative gene for type 3 RTA is located on the 8q22 locus and encodes, a protein called carbonic anhydrase II. In this study we analysed the entire exons and flanking regions of the gene CAII in one child suffering renal tubular acidosis with autosumal recessive pattern, and clinical diagnosis of type3 RTA. DNA was extracted from blood sample of patient by the salting out extraction method. The exons and flanking regions of the CAII gene were amplified using polymerase chain reaction (PCR). We performed exon direct sequencing by forward and reverse primers. No mutation was found following the screening of the entire coding sequence of the CAII gene. It is likely that another gene must be involved in this case. In other word another type of RTA must be considered

Autosomal dominant polycystic kidney disease (ADPKD) is one of the most common genetic kidney disorders with the incidence of 1 in 1،000 births. ADPKD is genetically heterogeneous with two genes identified: PKD1 (16p13. 3، 46 exons) and PKD2 (4q21، 15 exons). Eighty five percent of the patients with ADPKD have at least one mutation in the PKD1 gene. Genetic studies have demonstrated an important allelic variability among patients، but very few data are known about the genetic variation among Iranian populations. In this study، exon direct sequencing of PKD1 was performed in a seven-year old boy with ADPKD and in his parents. The patient’s father was ADPKD who was affected without any kidney dysfunction، and the patient’s mother was congenitally missing one kidney. Molecular genetic testing found a mutation in all three members of this family. It was a missense mutation GTG>ATG at position 3057 in exon 25 of PKD1. On the other hand، two novel missense mutations were reported just in the 7-year-old boy: ACA>GCA found in exon 15 at codon 2241 and CAC>AAC found in exon 38 at codon 3710. For checking the pathogenicity of these mutations، exons 15، 25، and 38 of 50 unrelated normal cases were sequenced. our findings suggested that GTG>ATG is a polymorphism with high frequency (60%) as well as ACA>GCA and CAC>AAC are polymorphisms with frequencies of 14% and 22%، respectively in the population of Southwest Iran.