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Avicenna Journal of Medical Biotechnology - Volume:8 Issue: 4, Oct-Dec 2016

Avicenna Journal of Medical Biotechnology
Volume:8 Issue: 4, Oct-Dec 2016

  • تاریخ انتشار: 1395/07/25
  • تعداد عناوین: 9
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  • Shahin Akhondzadeh Page 1
  • Maliheh Jahromi, Shahnaz Razavi, Nushin Amirpour, Zahra Khosravizadeh Page 152
    Background
    Some antidepressant drugs can promote neuronal cell proliferation in vitro as well as hippocampal neurogenesis in human and animal models. Furthermore, adipose tissue is an available source of adult stem cells with the ability to differentiate in to multiple lineages. Therefore, human Adipose-Derived Stem Cells (hADSCs) may be a suitable source for regenerative medical applications. Since there is no evidence for the effect of Paroxetine as the most commonly prescribed antidepressant drug for neurogenic potential of hADSCs, an attempt was made to determine the effect of Paroxetine on proliferation and neural differentiation of hADSCs.
    Methods
    ADSCs were isolated from human abdominal fat. These cells differentiated to neuron-like cells and were treated with Paroxetine. 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide (MTT) assay and immunofluorescence technique were used for assessment of cell proliferation and neurogenic differentiation potential of induced cells, respectively.
    Results
    MTT assay analysis showed that Paroxetine significantly increased the proliferation rate of induced hADSCs (p
    Conclusion
    Our results provide evidence that Paroxetine can promote proliferation and differentiation rate during neurogenic differentiation of hADSCs. Moreover, Paroxetine can reduce gliogenesis of induced hADSCs during neurogenic differentiation.
    Keywords: Antidepressant drugs, Neurogenic differentiation, Paroxetine, Proliferation, Stem cells
  • Ghamartaj Hossein, Manijeh Khanmohammadi, Parissa Sahranavard Fard, Yasaman Heidarian, Somaieh Kazemnejad, Mohammad Mehdi Akhondi Page 159
    Background
    It has been reported that secreted frizzled-related protein-4 known as an antagonist of Wnt signaling pathway plays a role in luteinization process of rodent granulosa cells. The purpose of this study was twofold: 1) to determine whether recombinant human secreted frizzled-related protein-4 (rhSFRP-4) could directly induce terminal differentiation of rat Granulosa Cells (GCs) and 2) to understand how the modulation of β-catenin and Protein Kinase B (PKB)/AKT activity by exogenous SFRP-4 could be involved in steroidogenesis.
    Methods
    GCs were firstly stimulated with Follicle-Stimulating Hormone (FSH) named as FSH-primed cells then were treated with luteinizing hormone (LH). Then estradiol (E2) and progesterone (P4) production levels were assessed in the absence or presence of rhSFRP-4 treatment. The expression levels of activated -catenin, pAKTser473, pGSK3ser9 were assessed by western blot or immuno-fluoresence.
    Results
    In the presence of rhSFRP-4, there was 38% decreased E2 levels compared to untreated FSH-primed cells (p
    Conclusion
    Taken together, our results showed that rhSFRP-4 could directly induce terminal differentiation in GCs via the modulation of β-catenin and PKB/AKT pathways and that it does so in a dose-dependent manner.
    Keywords: Active β catenin_GSK3β PKB_AKT_Rat granulosa cell_Secreted frizzled_related protein_4 (SFRP_4)
  • Elham Isaei, Shahla Mansouri, Fereshteh Mohammadi, Sadegh Taheritarigh, Zohreh Mohammadi Page 169
    Background
    Antibiotic resistant bacteria can be considered as a main problem in infection management. Zinc oxide nanoparticles (ZnO NPs), individually or in combination with antibiotics, can be considered as good candidates for struggling against drug resistant bacteria.
    Methods
    In this study, Zinc oxide nanoparticles were synthesized using sol-gel method in low temperature as a cost effective procedure and characterized by X-ray diffraction and Scanning Electron Microscopy. Antibacterial activity of 9 new combinations of Zinc oxide nanoparticles and ceftazidime was assessed against standards and new clinically isolated multi drug resistant Pseudomonas aeruginosa ( P. aeruginosa ), in order to evaluate enhancement effect of synthesized Zinc oxide nanoparticles on antibacterial activity of ceftazidime.
    Results
    The results indicated that desirable effects can be seen at 6 and 7 mM of Zinc oxide nanoparticles (60 to 100% inhibition). Moreover, after evaluation of 9 new combinations with various concentrations of both components, it was demonstrated that Zinc oxide nanoparticles can enhance the antibacterial activity of ceftazidime, against some bacterial strains of P. aeruginosa . The highest activity was observed with the concentration of 20 μ g/ml ceftazidime in the presence of 5, 6 or 7 mM of Zinc oxide nanoparticles.
    Conclusion
    Zinc oxide nanoparticles in appropriate concentrations can be proposed as new and promising candidates for overcoming bacterial resistance.
    Keywords: Antibiotic resistance, Ceftazidime, Pseudomonas aeruginosa, ZnO NPs
  • Asaad Azarnezhad, Zohreh Sharifi, Rahmatollah Seyedabadi, Arshad Hosseini, Behrooz Johari, Mahsa Sobhani Fard Page 175
    Background
    As a drug target and an antigenic agent, HIV-1 protease (HIV-1 PR) is at the center of attention for designing anti-AIDS inhibitors and diagnostic tests. In previous studies, the production of the recombinant protease has been faced with several difficulties; therefore, the aims of this study were the easy production, purification of the soluble form of protease in E. coli and investigation of its immunoreactivity.
    Methods
    Protease coding region was isolated from the serum of an infected individual, amplified by RT-PCR and cloned into PTZ57R using TA-cloning. Protease coding frame was isolated by PCR and cloned in pET102/D. TOPO expression vector and cloned protease was expressed in Escherichia coli ( E. coli) BL21 . Produced recombinant protein was purified by affinity Ni-NTA column and protein concentration was checked by BCA protein assay kit. Subsequently, immunoreactivity of recombinant protease (rPR) was assayed by Western blotting and ELISA.
    Results
    Cloning of the HIV protease by TOPO cloning system in pET102/D.TOPO was confirmed with PCR and sequencing. The concentration range of purified recombinant protein was 85 to 100  g/ml . Immunogenicity of rPR was confirmed by Western blotting and ELISA.
    Conclusion
    Soluble production of recombinant HIV-1 protease (HIV-1 rPR) was performed successfully. This recombinant protein disclosed 86% specificity and 90% sensitivity in immunoassay tests.
    Keywords: Human immunodeficiency virus, Molecular cloning, Protease, Recombinant proteins
  • Mozhdeh Zamani, Navid Nezafat, Younes Ghasemi Page 182
    Background
    In the recent years, there has been an increasing interest in secretory production of recombinant proteins, due to its various advantages compared with intracellular expression. Signal peptides play a critical role in prosperous secretion of recombinant proteins. Accordingly, different signal peptides have been assessed for their ability to produce secretory proteins by trial-and-error experiments. The aim of this study was to evaluate the effect of L-asparaginase II signal peptide on the recombinant human Growth Hormone (hGH) protein secretion in the Escherichia coli
    (E. coli) host.
    Methods
    Cloning and expression of a synthetic hGH gene, containing L-asparaginase II signal sequence was performed in E. coli BL21 (DE3) using 0.1mM IPTG as an inducer at 23 C overnight . Periplasmic protein extraction was performed using three methods, including osmotic shock, osmotic shock in the presence of glycine and combined Lysozyme/EDTA osmotic shock. Afterwards, the hGH expression was determined by SDSPAGE.
    Results
    Based on experimentally obtained results, hGH protein is expressed as inclusion body even in the presence of L-asparaginase II signal peptide.
    Conclusion
    Therefore, this signal peptide is not effective for secretory production of the recombinant hGH.
    Keywords: Escherichia coli (E. coli), Human growth hormone, L, asparaginase II, Recombinant proteins, Signal peptide
  • Tahereh Sadeghiyan Rizi, Jamshid Fooladi, Sima Sadrai Page 188
    Background
    L-tryptophan is used widespread in the pharmaceutical industry. The majority of L-Trp production depends on microbial processes that produce L-tryptophan from indole and L-serine. These processes are very costly due to the costs of precursors, especially L-serine. Use of inexpensive substitutions as the L-serine source of Ltryptophan production enables us to reach a cost-effective process. In this paper, effect of Triton X-100 on L-Trp production and the ability to use Iranian cane molasses as inexpensive L-serine source was investigated.
    Methods
    Escherichia coli (E. coli) ATCC 11303 cells were grown in 10-L fermenter containing minimal medium supplemented with beet molasses as an inexpensive carbon source and indole as tryptophan synthase inducer. Whole cells of stationary phase were used as biocatalyst for L-Trp production. Triton X-100 addition to the production medium as indole reservoir was investigated. Then, cane molasses was used as LSer source in L-Trp production medium. Amount of L-Tryptophan and theoretical yield of L-Trp production was determined by HPLC and by a colorimetrically method on the basis of remaining indole assay, respectively.
    Results
    As a result, triton X-100 increased L-Trp production three times. Also, the result showed that 0.68 mM L-Tryptophan was produced in the presence of cane molasses at 37 C for 8 hr .
    Conclusion
    This result showed that cane molasses of Qazvin sugar factory includes significant amounts of L-Ser that makes it a suitable substitution for L-Ser in L-Trp production. Therefore, it has the potential to be used for cost-effective L-Trp production in industrial scale.
    Keywords: Molasses, PLP, Tryptophan synthase, Tryptophanase
  • Zahra Ghasemi, Mehrdad Hashemi, Mahsa Ejabati, Seyyed Meisam Ebrahimi, Hamidreza Kheiri Manjili, Ali Sharafi, Ali Ramazani Page 193
    Background
    Genetic polymorphisms of drug metabolisms by cytochrome P450 (P450s) could affect drug response, attracting particular interest in the pharmacogenetics. Due to the importance of CYP2C19* 17 allele and its capability of super- fast metabolism and also lack of information about distribution of the alleles in Iranian population, this research aimed to use High Resolution Melting (HRM) method compared to PCR-RFLP for genotyping healthy Iranian population.
    Methods
    Blood samples were collected from 100 healthy Iranian volunteers. DNA was extracted by salting out method. Real-time PCR was used for amplification of the CYP2C19 gene and the alleles were identified by HRM. Sequencing was used to confirm the amplified DNA fragments and data were analyzed using SPSS software ver.18.
    Results
    The frequency of alleles CYP2C19*1/*1, CYP2C19*1/*17 and CYP2C19*17/*17 were estimated as 58.33, 29.1 and 11.1%, respectively. Specificity and sensitivity of HRM method were 90% and 100%, with respect to PCR-RFLP. Also, HRM analysis has been evaluated as a faster and more effective approach.
    Conclusion
    Comparison of our results based on HRM analysis with PCR-RFLP showed that our developed method is rapid, accurate, fast and economic to study the CYP2C19*17 allele and it is appropriate for other similar population genetic studies.
    Keywords: Cytochrome P, 450 CYP2C19, Pharmacogenetics, Real, Time Polymerase Chain Reaction
  • Isaac Firouze Moqadam, Shamsolmoulouk Najafi, Mahsa Mohammadzadeh, Alireza Zare Bidoki, Hila Yousefi, Elham Farhadi, Arghavan Tonekaboni, Ghasem Meighani, Mohsen Mohammadzadeh, Ali Akbar Amirzargar, Nima Rezaei Page 200