1 17286158 4 100 Stable Transfection of pEGFP-N1-MOG Plasmid to Utilize in Multiple Sclerosis Gene Therapy Karimfar Mohammad Hassan b Noorozian Mohsen c Mastery Farahani Reza d Sadat Khoramgah Maryam e Azimi Hadi f Keramatinia Ali Asghar g Ahmadi Rozbahani Navid h Bahadori Monfared Ayad i Abbaszadeh Hojjat-Allah j b Organization c Organization d Organization e Organization f Organization g Organization h Organization i Organization j Department of Biology and Anatomical Sciences, Shahid Beheshti University of Medical Sciences and Health Services, Tehran, Iran. 1 2 2015 12 1 3 8 18 08 2014 02 12 2014 Introduction: Multiple sclerosis (MS) is a disease of the immune system: it attacks the myelin around the axons and leaves them exposed. Destruction of myelin weakens the electrical conduction of ions and thus leads to a lack of communication in the nervous system.  Methods: In the present study, we constructed recombinant plasmid and then transformed to E. coli cell. The colonies containing plasmid were selected by Colony PCR. Enzyme digestion and sequencing were utilized to approve the accuracy of the extracted plasmid of these clones. Recombinant plasmid transfect in to mesanchymal stem cells. Results: Plasmid was verified correctly. After transfection, the transcription of MOG gene and the expression of MOG protein were proved by RT-PCR, western blotting and Elisa. Conclusion: Plasmid was constructed correctly and mesenchyme stem cells were successfully transfected by transfection and protein can be expressed well, setting a proper foundation for the future studies on the transplantation of gene modified mesanchymal stem cells in order to promote Multiple sclerosis.
101 Association of rs12913832 in the HERC2 Gene Affecting Human Iris Color Variation Rafati Alireza k Hosseini Mostafa l Tavallaei Mahmoud m Naderi Mostafa n Mohammadi Amir Hashem o Bahmani Hasan p Sarveazad Arash k Organization l Department of Medical Genetics, Human Genetic Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran. m Organization n Organization o Organization p Organization Organization 1 2 2015 12 1 9 16 06 07 2014 23 10 2014 Introduction: Human eye colour as a physical trait is based on the developmental biology and genetic determinants of the structure known as the iris, which is part of the uveal tract of the eye. Prediction of human visible characteristics (EVCs) by genotyping informative SNPs in DNA as biological witness opens up a new avenue in the forensic genetic. Variation of iris color rely on the amounts of eumelanine and pheomelanin. The aim of this research was to determine and evaluate the frequency and the association of rs12913832 with prediction of human eye color in 53 volunteer of Iranian population samples. Methods: A selection of human body blood samples were collected from donors with informed consent in Clinic Ophthalmology of Baqiyatallah hospital. DNA was extracted from the samples using RGDE procedure. PCR primers for rs12913832 were designed to give amplicon sizes up to 189 bp and Single base extensions (SBE) were done by applying the SNaPshot Multiplex kit in 6 ;mul reaction volumes. The results were analyzed with the SPSS 22.0 software package. Results: The frequency of eye color were achieved for brown 34%, blue 17% and intermediate colors 49%, respectively. The genotype frequencies of T/T, C/T and C/C in our population were 4.26 %, 8.35 % and 7.37%, respectively. The statistical analysis revealed the two genotypes including T/T and C/C had a significant associate with dark brown eyes and bright blue eyes, respectively. The sensitivity and specificity of our method were determined 100% and 56.25%,respectively. Conclusion: Our results demonstrated that rs12913832 C>T polymorphism is associated with blue iris color in Iranian population. However, assessment SNP markers by using SNaPshot is a key tool for tracing unknown persons to get primarily information about genotypic and phenotypic characteristics. 112 The Effects of Iron Oxide Nanoparticle on Differentiation of Human Mesenchymal Stem Cells to Osteoblast Foroutan Tahereh Zaman Kasaie Mohamad Organization Department of Animal Biology, Faculty of Biological Sciences, Kharazmi University, Tehran, Iran. 1 2 2015 12 1 17 22 23 09 2014 05 01 2015 Introduction: IIron oxide nanoparticles (IO NP) have an increasing number of biomedical applications. To date, the potential cytotoxicity of these particles remains an issue of debate. Little is known about the cellular interaction or toxic effects of IO NP on differentiation of stem cells. The aim of the present study was to investigate the possible toxic role of different doses of IO NP in differentiation of human mesenchymal stem cells (hMSCs) derived bone marrow to osteoblast. Methods: hMSCs were seeded on normal stem cell medium with added 20 and 70µg/ml of IO NPs. Post confluence (2 weeks), cells growing cell viability was measured by 3-(4, 5-Dimethylthiazol-2-yl) -2, 5 -diphenyltetrazolium bromide (MTT) assays. hMSCs at passage 2 were cultured in osteogenic medium added with 20 and 70 µg/ml of IO NPs. The expression of osteogenesis markers of osteopontin, osteocalcine in different groups was compared by RT-PCR assay. Results: Our findings showed that cell viability of hMSCs cultured in normal stem cell media containing 20µg/ml IO NPs was significantly (P>0.05) lower than control and 70µg/ml dose of IO NPs groups, while there was no significant difference between control and 70µg/ml dose of IO NP. The expression level of osteoblast markers osteopontin and osteocalcin in hMSCs differentiated to osteoblast in dose with 20µg/ml IO NPs was significantly lower than the other groups, while the expression level of osteopontin and osteocalcin in 70µg/ml IO NPs dose was insignificantly higher than control group. Conclusion: To summarize, the presence of IO NPs with low dose influenced the cell viability cells in normal stem cell media, demonstrating toxicity of this material with 20µg/ml. It could be probably due to penetrating particles throughout cell membrane. This represents a critical aspect to its successful use for stem cell-based regenerative medicine strategies. 89 Establishment of Oxidative Stress Modeling during Spermatogonial Stem Cells Cultivation Treated with Different Doses of H2O2 Barati Shirin Movahedin Mansoureh Mazaheri Zohreh Batooli Hosein Tahmasebi Fateme Tarbiat Modares University, Faculty of Medical Sciences, Department of Anatomical Sciences. Kashan Botonical Garden, Isfahan Research Center of Agriculture and Natural Resources, Isfahan, Iran. Tarbiat Modares University, Faculty of Medical Sciences, Department of Anatomical Sciences. Kashan Botonical Garden, Isfahan Research Center of Agriculture and Natural Resources Tarbiat Modares University, Faculty of Medical Sciences, Department of Anatomical Sciences. 1 2 2015 12 1 23 28 12 09 2014 14 11 2014 Introduction: Nowadays, spermatogonial stem cells (SSCs) cultivation has been used by many researchers as an effective tool for infertility treatments. Oxidative conditions can be effective on cell proliferation and differentiation of these cells. So, the aim of this study was to establish oxidative stress model for antioxidant activity of some drugs investigation during SSCs in vitro culture. Methods: Neonatal NMRI male mice (3-5 day) were used for isolation of SSCs. The cell suspension was prepared by twice enzymatic digestion. The cell suspension contents were spermatogonial and sertoli cells and treated by different doses of H2O2 logarithmic concentrations from 0-100 ;muM after 24 hours. To access the optimal dose, extra doses from 10-100;muM was evaluated. After 2 hours of H2O2 treatment, viability was determined by Trypan blue assay. The data were analyzed using SPSS software and One-way ANOVA test. Results: Our data showed that spermatogonial stem cells colonies appeared after 4 days of isolation. These cells expressed OCT4 and PLZF proteins. Many of spermatogonial stem cells were removed after using higher doses of H2O2. The results showed that 30 ;muM concentration of H2O2 could induce oxidative stress in spermatogonial stem cell during in vitro culture. Conclusion: According to this study, 30 ;muM concentration of H2O2 can cause cell death lower than 50% of total number of cells and can increase oxidative stress in cultivation of SSCs. This model is a suitable tool for studying some new antioxidant drugs. 81 Studying of the Expression of BAX and BCL-2 Genes in Men with Sperm DNA Fragmentation Based on DFI and Comet Assay Taheri Hossein Salehi Mohammad Eslami Arshaghi Tarlan Mosahebi Majid Organization Cellular and Molecular Biology Research Center, Shahid Beheshti University of Medical Sciences and Health Services, Tehran, Iran. Organization Organization 1 2 2015 12 1 29 36 01 07 2014 28 10 2014 Introduction: The amount of expression of BAX and BCL-2 genes in infertile men’s sperm as well as its association with sperm parameters and DNA fragmentation index is an issue which has not been studied yet. In this research, it is assumed that up-regulation of BAX and downregulation of BCL-2 are directly associated with sperm DNA fragmentation. Methods: After obtaining semen samples from the patients by using gradient centrifugation method, the semen samples were centrifuged using gradient method in order to obtain pure sperm. Sperm is divided into three parts based on which flow cytometry, Real Time-PCR and Comet Assay techniques were conducted. After extracting RNA and producing cDNA, the amount of expression of BAX and BCL-2 genes was measured using Real Time-PCR. The amount of sperm DNA fragmentation was measured using flow cytometry and Comet Assay techniques. Based on the amount of DNA fragmentation index (DFI), samples were divided into the two groups of control (DFI<30) and DNA fragmentation (DFI;ge30). Using WHO criteria, sperm parameters (morphology and motility) were evaluated. Results: This study showed that the amount of expression of BAX in the DNA fragmentation group was not significant compared to the control group but the expression of BCL-2 gene decreased significantly (P<0.05). Also, in many cases, there was a significant difference between the two groups in terms of parameters of sperm DNA fragmentation and morphological parameters (P<0.05). Conclusion: This study showed that reduction of expression of BCL-2 increases sperm DNA damage and this result can be helpful for therapeutic purposes. 103 Computed Tomographic Anatomy of the Abdominal Cavity in the Jebeer (Gazella Bennettii) Sajjadian Seyed Mohsen Shojaei Bahador Sohrab Zade Babak Department of Basic Sciences, Faculty of Veterinary Medicine, Shahid Bahonar University of Kerman, Kerman, Iran. Department of Basic Sciences, Faculty of Veterinary Medicine, Shahid Bahonar University of Kerman, Kerman, Iran. Doctor of the Veterinary Medicine 1 2 2015 12 1 37 44 21 08 2014 10 12 2014 Introduction: Computed tomography (CT) is an imaging technique with gives us an opportunity to review cross sections of the body in live animals. In veterinary medicine although CT is mostly used for diagnostic purposes in small animals, but in recent years CT also has been used as a non invasive modality in non clinical studies. Jebeer Gazelle (Gazella bennettii) is one of the species of the genus Gazella which lives mostly in the south east of Iran and very little anatomic studies are available on it. The aim of this study was preparing detailed anatomic images of the abdominal cavity of the Jebeer as an endangered species using the non invasive CT technique. Methods: Spiral CT images were acquired from the abdominal region of four healthy Jebeer, perpendicular to long axis of the body. CT windows were adjusted as necessary to have optimized images of the abdominal organs. The images were studied serially and compared anatomically with two dissected goats. Results: Liver, spleen, reticulum, omasum, abomasum, rumen, right and left kidneys, transverse colon, ascending colon, descending colon, cecum, pancreas, duodenum, uterine horns, urinary bladder and jejunum were distinguished and addressed according to the thoracic and lumbar vertebrae as landmarks. Conclusion: According to the present study, we identified the abdominal organs, their precise position and related structures in the Jebeer without any invasive procedure. This is the first study which is addressing abdominal organs of a wild ruminant by using CT modality. 113 Case Report: Congenitally Missing Teeth Jahanimoghadam Fatemeh Torabi Moolok Rostami Shima Dental School, Department of Pediatric Dentistry, Oral & Dental Research Center, Kerman University of Medical Sciences, Kerman, Iran. Organization Organization 1 2 2015 12 1 45 50 11 08 2014 29 11 2014 Congenitally missing of maxillary lateral incisors is one of the most common patterns of hypodontia. This paper presents a nine year old boy with congenital missing of lateral incisors. Familial history showed that, his mother, aunts, uncle and grandmother have also congenital absence of lateral incisors. 85 Case Report: Variation of the Gallbladder Vasculature Including Double Cystic Arteries Eyni Hossein Pasbakhsh Parichehr Azimi Arian Behzadi Fard Saba Tarbiat Modares University Department of Anatomy, School of Medicine, University of Tehran, Tehran, Iran. Tarbiat Modares University Tarbiat Modares University 1 2 2015 12 1 51 55 16 11 2014 07 01 2015 Acquaintance with the different anatomical variations of the arterial supply of the gallbladder is of great importance in hepatobiliary surgical procedures. A rare variation of the hepatobiliary arterial system was found during anatomical dissection of a female Iranian cadaver. Two cystic arteries were present, the first arising directly from the right hepatic artery and the second from the gastroduodenal artery. Knowledge of the different anatomical variations of the arterial supply of the gallbladder is of great importance in hepatobiliary surgical procedures.