Avicenna Research InstituteAvicenna Journal of Medical Biotechnology2008-28358420161001Paroxetine Can Enhance Neurogenesis During Neurogenic Differentiation of Human Adipose-derived Stem Cells152158250190ENMalihehJahromiDepartment of Anatomical Sciences, Faculty of Medicine, Isfahan University of Medical Sciences, Isfahan, IranShahnazRazaviDepartment of Anatomical Sciences, Faculty of Medicine, Isfahan University of Medical Sciences, Isfahan, IranNushinAmirpourDepartment of Anatomical Sciences, Faculty of Medicine, Isfahan University of Medical Sciences, Isfahan, IranZahraKhosravizadehDepartment of Anatomical Sciences and Molecular Biology, Faculty of Medicine, Tehran University of Medical Sciences, Tehran, IranJournal Article20151206Background: 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<0.05), while immunofluorescent staining indicated that Paroxetine treatment during neurogenic differentiation could enhance the mean percentage of Nestin and MAP2 (Microtubule-associated protein-2) positive cells but the mean percentage of GFAP (Glial acidic fibrillary protein) positive cells significantly decreased relative to control group (p<0.05). 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.https://www.ajmb.org/article_250190_9f4a0a37e62e884dd90a9e8c256c0722.pdfAvicenna Research InstituteAvicenna Journal of Medical Biotechnology2008-28358420161001Exogenous Secreted Frizzled-Related Protein-4 Modulates Steroidogenesis of Rat Granulosa Cells Through Wnt/Bcatenin and PI3K/AKT Signaling Pathways159168250191ENGhamartajHosseinDepartment of Animal Physiology, Developmental Biology Laboratory, Faculty of Biology, University College of Science, University of Tehran, Tehran, IranManijehKhanmohammadiDepartment of Animal Physiology, Developmental Biology Laboratory, Faculty of Biology, University College of Science, University of Tehran, Tehran, Iran; Reproductive Biotechnology Research Center, Avicenna Research Institute, ACEParissaSahranavard FardDepartment of Animal Physiology, Developmental Biology Laboratory, Faculty of Biology, University College of Science, University of Tehran, Tehran, Iran; Department of Stem Cells & Developmental Biology at Cell Science Research CeYasamanHeidarianDepartment of Animal Physiology, Developmental Biology Laboratory, Faculty of Biology, University College of Science, University of Tehran, Tehran, IranSomaiehKazemnejadReproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, Iran0000-0003-1808-8357Mohammad MehdiAkhondiReproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, IranJournal Article20160410Background: 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 (P < sub>4) production levels were assessed in the absence or presence of rhSFRP-4 treatment. The expression levels of activated β-catenin, pAKTser 473 , pGSK3βser 9 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<0.05), and P < sub>4 production subsequently decreased. However, in GCs pre-treated with rhSFRP-4 prior to addition of FSH, P < sub>4 levels increased 2-fold compared with untreated cells (p<0.05). Unexpectedly, treatment with rhSFRP-4 prior to LH stimulation inhibited LH-induced P < sub>4 secretion. Treatment with low (0.5 ng/ml) but not high (50 ng/ml) concentrations of rhSFRP-4 led to significantly increased levels of pGSK3βser 9 (1.6-fold) and nuclear active β-catenin (2.8-fold) in GCs compared with untreated cells. Interestingly, pre-treating GCs with rhsFPR4 prior to LH stimulation resulted in a 38% decrease in pAKTser 473 levels compared with those in LH-treated cells (p<0.05). 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.https://www.ajmb.org/article_250191_c6d174062a70a908e8d981e35670a853.pdfAvicenna Research InstituteAvicenna Journal of Medical Biotechnology2008-28358420161001Novel Combinations of Synthesized ZnO NPs and Ceftazidime: Evaluation of their Activity Against Standards and New Clinically Isolated Pseudomonas aeruginosa169174250192ENElhamIsaeiDepartment of Microbiology, Afzalipour Faculty of Medicine, Kerman University of Medical Sciences, Kerman, IranShahlaMansouriDepartment of Microbiology, Afzalipour Faculty of Medicine, Kerman University of Medical Sciences, Kerman, IranFereshtehMohammadiNanobiotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, IranSadeghTaheritarighDepartment of Plant Breeding and Biotechnology, Faculty of Plant Production, Gorgan University of Agricultural Sciences and Natural Resources, Gorgan, IranZohrehMohammadiNanobiotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, IranJournal Article20160213Background: Antibiotic resistant bacteria can be considered as a main problem in infection management. Zinc oxide nanopajmb_articles (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 nanopajmb_articles 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 nanopajmb_articles 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 nanopajmb_articles on antibacterial activity of ceftazidime. Results: The results indicated that desirable effects can be seen at 6 and 7 mM of Zinc oxide nanopajmb_articles (60 to 100% inhibition). Moreover, after evaluation of 9 new combinations with various concentrations of both components, it was demonstrated that Zinc oxide nanopajmb_articles can enhance the antibacterial activity of ceftazidime, against some bacterial strains of P. aeruginosa. The highest activity was observed with the concentration of 20 μg/ml ceftazidime in the presence of 5, 6 or 7 mM of Zinc oxide nanopajmb_articles. Conclusion: Zinc oxide nanopajmb_articles in appropriate concentrations can be proposed as new and promising candidates for overcoming bacterial resistance.https://www.ajmb.org/article_250192_fe96a2179425a979d1427a2e863a1f42.pdfAvicenna Research InstituteAvicenna Journal of Medical Biotechnology2008-28358420161001Cloning and Expression of Soluble Recombinant HIV-1 CRF35 Protease-HP Thioredoxin Fusion Protein175181250193ENAsaadAzarnezhadCellular and Molecular Research Center, Kurdistan University of Medical Sciences, Sanandaj, Iran; Department of Medical Genetics, Faculty of Medicine, Tehran University of Medical Science, Tehran, IranZohrehSharifiBlood Transfusion Research Center, Institute for Research and Education in Transfusion Medicine, Tehran, IranRahmatollahSeyedabadiDepartment of Molecular Medicine and Genetics, Faculty of Medicine, Hamedan University of Medical Sciences, Hamedan, IranArshadHosseiniDepartment of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences, Tehran, IranBehroozJohariDepartment of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences, Tehran, IranMahsaSobhani FardDepartment of Immunology, Faculty of Medicine, Hamedan University of Medical Sciences, Hamedan, Iran; Student Research Center, Hamedan University of Medical Sciences, Hamedan, IranJournal Article20160118Background: 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 μg/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.https://www.ajmb.org/article_250193_1c52048e9ce52de3a89d0b037d333d8b.pdfAvicenna Research InstituteAvicenna Journal of Medical Biotechnology2008-28358420161001Evaluation of Recombinant Human Growth Hormone Secretion in E. coli using the L-asparaginase II signal Peptide182187250194ENMozhdehZamaniPharmaceutical Sciences Research Center, Shiraz University of Medical Sciences, Shiraz, Iran; Department of Pharmaceutical Biotechnology, Faculty of Pharmacy, Shiraz University of Medical Sciences, Shiraz, IranNavidNezafatPharmaceutical Sciences Research Center, Shiraz University of Medical Sciences, Shiraz, IranYounesGhasemiPharmaceutical Sciences Research Center, Shiraz University of Medical Sciences, Shiraz, Iran; Department of Pharmaceutical Biotechnology, Faculty of Pharmacy, Shiraz University of Medical Sciences, Shiraz, IranJournal Article20151229Background: 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 0.1 mM IPTG as an inducer at 23 o 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 SDS-PAGE. 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.https://www.ajmb.org/article_250194_bf14357628acc81e74c7f9ebdc08fdb1.pdfAvicenna Research InstituteAvicenna Journal of Medical Biotechnology2008-28358420161001Preliminary Study on Cost-effective L-Tryptophan Production from Indole and L-Serine by E. coli Cells188192250195ENTaherehSadeghiyan-RiziDepartment of Pharmaceutical Biotechnology, Faculty of Pharmacy, Isfahan University of Medical Sciences, Isfahan, IranJamshidFooladiNational Laboratory of Industrial Microbiology, Department of Biology, Faculty of Science, Alzahra University, Tehran, IranSimaSadraiDepartment of Pharmaceutics, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran, IranJournal Article20160106Background: 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 L-tryptophan 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 L-Ser 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 o 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.https://www.ajmb.org/article_250195_cbf80df28f807e85b5c73765ee69d942.pdfAvicenna Research InstituteAvicenna Journal of Medical Biotechnology2008-28358420161001Development of a High-Resolution Melting Analysis Method for CYP2C19*17 Genotyping in Healthy Volunteers193199250196ENZahraGhasemiDepartment of Genetics, Tehran Medical Sciences Branch, Islamic Azad University, Tehran, IranMehrdadHashemiDepartment of Genetics, Tehran Medical Sciences Branch, Islamic Azad University, Tehran, IranMahsaEjabatiDepartment of Biotechnology, Faculty of Pharmacy, Zanjan University of Medical Sciences, Zanjan, IranSeyyed MeisamEbrahimiDepartment of Medical Surgical Nursing, Abhar Faculty of Nursing, Zanjan University of Medical Sciences, Zanjan, IranHamidrezaKheiri ManjiliDepartment of Biotechnology, Faculty of Pharmacy, Zanjan University of Medical Sciences, Zanjan, IranAliSharafiDepartment of Biotechnology, Faculty of Pharmacy, Zanjan University of Medical Sciences, Zanjan, IranAliRamazaniDepartment of Biotechnology, Faculty of Pharmacy, Zanjan University of Medical Sciences, Zanjan, IranJournal Article20160120Background: Genetic polymorphisms of drug metabolisms by cytochrome P450 (P450s) could affect drug response, attracting particular interest in the pharm-acogenetics. 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.https://www.ajmb.org/article_250196_62c29fbf3d4e7aef7b074f6c4d726d1a.pdf