Depression and Inflammation: Is There any Role for Biomarkers? 2 2 Despite the advent of several antidepressant medications, treatment of Major Depressive Disorder (MDD) is still far from optimal 1-3.  A large proportion of patients with MDD do not respond to their first medication. To achieve favorable response, these patients are generally treated with either switching to another treatment or augmentation therapy. In the recent decade, several augmentative strategies for treatment of MDD have been developed. Some of these treatment modalities focus on recently developed hypotheses of pathophysiological processes in patients with MDD 1-3. These mainly include immune system dysfunction, hypothalamic-pituitary-adrenal (HPA) axis and metabolic derangements, impaired neuroprotection, or neuroinflammation 1-3. Growing body of evidence suggests that inflammation is implicated in the pathophysiology of MDD 4-6. Sickness be-havior which is a result of inflammatory activation, shares many clinical features such as anhedonia, anorexia, irritability, and mild cognitive problems with MDD 4-6. Several studies have shown an elevation of proinflammatory cytokines [particularly IL-6 and Tumor Necrosis Factor (TNF-α)] in patients with MDD 7. A large body of research now suggests that depression is associated with a low-grade, chronic inflammatory response and is accompanied by increased oxidative stress.  • depression frequently is comorbid with many inflammatory illnesses  • increased inflammatory biomarkers are associated with major depressive disorder (MDD)  • exposure to immunomodulating agents may increase the risk of developing depression  • stress can activate proinflammatory pathways  • antidepressants can decrease inflammatory response  • inhibition of inflammatory pathways can improve mood 4-7. IL-6 is one of the most widely studied cytokines in patients with MDD 8,9. In addition to elevation of this cytokine in patients with MDD, relation of IL-6 concentration to severity of depression, a shift in circadian rhythm 8,9, and a reduction in its concentration in response to antidepressants have been shown in several studies.   Previous studies have already shown that elevated levels of inflammation are associated with poor response to antidepressants. The scientists found that they could pinpoint a threshold and precisely predict which patients would respond to conventional antidepressants. None of the patients with MIF and IL-1β levels above the threshold responded to the antidepressants most often prescribed. Those with inflammation levels below the threshold would likely respond. One reason for the lack of predictive biomarkers in MDD is that little is known with absolute certainty about how antidepressants improve mood. All currently approved medications for depression act in a similar way, increasing the availability of monoamine neurotransmitters like serotonin in the brain. Psychiatrists continue to search for biomarkers to help guide therapy and, potentially, improve chances of discovering new drugs 9.  In conclusion, the link between depression and the body's inflammatory response continues getting stronger, with more research showing an ever-tighter correlation. 207 207 Shahin Akhondzadeh Psychiatric Research Center, Roozbeh Hospital, Tehran University of Medical Sciences Psychiatric Research Center, Roozbeh Hospital, Tehran University of Medical Sciences Iran Editorial 20401.pdf Zeinoddini A, Sorayani M, Hassanzadeh E, Arbabi M, Farokhnia M, Salimi S, et al. Pioglitazone adjunctive therapy for depressive episode of bipolar disorder: a randomized, double-blind, placebo-controlled trial. Depress Anxiety 2015;32(3):167-173. ##Kashani L, Raisi F, Saroukhani S, Sohrabi H, Modabbernia A, Nasehi AA, et al. Saffron for treatment of fluoxetine-induced sexual dysfunction in women: randomized double-blind placebo-controlled study. Hum Psychopharmacol 2013;28(1):54-60.##Khajavi D, Farokhnia M, Modabbernia A, Ashrafi M, Abbasi SH, Tabrizi M, et al. Oral scopolamine augmentation in moderate to severe major depressive disorder: a randomized, double-blind, placebo-controlled study. J Clin Psychiatry 2012;73(11):1428-1433.##Akhondzadeh S, Jafari S, Raisi F, Nasehi AA, Ghoreishi A, Salehi B, et al. Clinical trial of adjunctive celecoxib treatment in patients with major depression: a double blind and placebo controlled trial. Depress Anxiety 2009;26(7):607-611.##Jeon SW, Kim YK. Inflammation-induced depression: Its pathophysiology and therapeutic implications. J Neuroimmunol 2017;313:92-98.##Pariante CM. Why are depressed patients inflamed? A reflection on 20 years of research on depression, glucocorticoid resistance and inflammation. Eur Neuropsychopharmacol 2017;27(6):554-559.##Dantzer R, O'Connor JC, Freund GG, Johnson RW, Kelley KW. From inflammation to sickness and depression: when the immune system subjugates the brain. Nat Rev Neurosci 2008;9(1):46-56.##Leighton SP, Nerurkar L, Krishnadas R, Johnman C, Graham GJ, Cavanagh J. Chemokines in depression in health and in inflammatory illness: a systematic review and meta-analysis. Mol Psychiatry 2018;23(1):48-58.##Abbasi SH, Hosseini F, Modabbernia A, Ashrafi M, Akhondzadeh S. Effect of celecoxib add-on treatment on symptoms and serum IL-6 concentrations in patients with major depressive disorder: randomized double-blind placebo-controlled study. J Affect Disord 2012;141(2-3):308-314.##

The p53 Modulated Cytotoxicity of Ophiocoma scolopendrina Polysaccharide Against Resistance Ovarian Cancer Cells 2 2 Background: Marine environment is a valuable source of bioactive compounds with variable medicinal properties. Previously, it was shown that Ophiocoma erinaceus extracted polysaccharide has prominent cytotoxic effect on HeLa human cervical cancer cells. In the present study, the anti-cancer properties of polysaccharide extracted from Ophiocoma scolopendrina (O. scolopendrina) were examined in comparison with paclitaxel as a conventional drug against resistant ovarian cancer; also, its related mechanism against A2780cp ovarian cancer cells was investigated.  Methods: The A2780cp cancer cells and NIH3T3 normal cells were cultured and treated with different concentrations of polysaccharide extracted from O. scolopendrina for 24 hr and 48 hr. Then, cell toxicity was studied by MTT assay, morphology of cells was observed under inverted microscopy and the type of induced cancer cell death was assessed by annexin V-FITC, propodium iodide and acridine orange staining. Finally, the apoptosis pathway was determined by measurement of caspase-3 and caspase-9 activity and assessment of p53 and Bcl-2. The statistical analysis was performed by SPSS software, one way ANOVA and p<0.05 was considered significant.  Results: Our observations from MTT assay and morphological assessment exhibited that O. scolopendrina isolated polysaccharide inhibited proliferation of ovarian cancer cells with IC50 of 35 µg/ml, while paclitaxel suppressed tumor cell growth with IC50=10 µg/ml. In contrast, MTT observations revealed low cytotoxicity of these chemotherapeutic agents against NIH3T3 normal cells. Also, the analysis correlated with induced cell death elucidated that concurrent treatment of polysaccharide plus paclitaxel had a further anti-cancer effect against A2780cp cells mainly through restoration of p53 and mitochondrial apoptosis cell death induction.  Conclusion: Taken together, our research supports the finding that application of polysaccharide extracted from O. scolopendrina can be considered a promising marine chemotherapeutic approach for advancing efficacy of paclitaxel in treatment of resistant ovarian cancer. Additional in vivo experiments are required to elucidate the role of brittle star polysaccharides in animal and clinical trials. 208 214 Elaheh Amini Department of Cellular & Molecular Biology, Faculty of Biological Sciences, Kharazmi University Department of Cellular & Molecular Biology, Faculty of Biological Sciences, Kharazmi University Iran Javad Baharara Department of Biology, Research Center for Animal Development Applied Biology, Mashhad Branch, Islamic Azad University Department of Biology, Research Center for Animal Development Applied Biology, Mashhad Branch, Islamic Azad University Iran Mahbube Afzali Department of Biology, Mashhad Branch, Islamic Azad University Department of Biology, Mashhad Branch, Islamic Azad University Iran Najme Nikdel Department of Biology, Mashhad Branch, Islamic Azad University Department of Biology, Mashhad Branch, Islamic Azad University Iran ApoptosisOvarian neoplasmsPaclitaxel 10376.pdf Mutee AF, Salhimi SM, Ghazali FC, Al-Hassan FM, Lim CP, Ibrahim K, et al. Apoptosis induced in human breast cancer cell line by Acanthaster planci starfish extract compared to tamoxifen. African J Pharm Pharmacol 2012;6(3):129-134.##Ooi KL, Muhammad TS, Sulaiman SF. Growth arrest and induction of apoptotic and non-apoptotic programmed cell death by, Physalis minima L. chloroform extract in human ovarian carcinoma Caov-3 cells. J Ethnopharmacol 2010;128(1):92-99.##Alizadehnohi M, Nabiuni M, Nazari Z. The lethal effect of honey bee venom on human ovarian cancer cisplatin resistance cell line A2780cp. Int Conf Environ Biomed Biotechnol 2012;41:46-49.##Kumaran NS, Bragadeeswaran S, Thangaraj S. Antimicrobial activities in star fishes Protoreaster lincki (Blainville, 1830) and Pentaceraster regulus (Muller & Troschel, 1842) againstisolated humafish pathogenic and biofilm microorganisms. J Appl Sci Res 2011;7(6):818-825.##Tai J, Cheung S, Wu M, Hasman D. Antiproliferation effect of Rosemary (Rosmarinus officinalis) on human ovarian cancer cells in vitro. Phytomedicine 2012;19(5):436-443.##Meshkini A, Yazdanparast R. Involvement of oxidative stress in taxol-induced apoptosis in chronic myelogenous leukemia K562 cells. Exp Toxicol Pathol 2012;64(4):357-365.##Fulda S. Modulation of apoptosis by natural products for cancer therapy. Planta Med 2010;76(11):1075-1079.##Folmer F, Jaspars M, Schumacher M, Dicato M, Diederich M. Marine natural products targeting phospholipases A2. Biochem Pharmacol 2010;80(12):1793-1800.##Senni K, Pereira J, Gueniche F, Delbarre-Ladrat C, Sinquin C, Ratiskol J, et al. Marine polysaccharides: a source of bioactive molecules for cell therapy and tissue engineering. Mar Drugs 2011;9(9):1664-1681.##Venkatesan J, Bhatnagar I, Kim SK. Chitosan-alginate biocomposite containing fucoidan for bone tissue engineering. Mar Drugs 2014;12(1):300-316.##Zong A, Cao H, Wang F. Anticancer polysaccharides from natural resources: a review of recent research. Carbohydr Polym 2012;90(4):1395-1410.##Angelova S, Gospodinova Z, Krasteva M, Antov G, Lozanov V, Markov T, Bozhanov S, et al. Antitumor activity of Bulgarian herb Tribulus terrestris L. on human breast cancer cells. J BioSci Biotech 2013;2(1):25-32.##Ramasamy S, Abdul Wahab N, Zainal Abidin N, Manickam S. Effect of extracts from Phyllanthus watsonii Airy Shaw on cell apoptosis in cultured human breast cancer MCF-7 cells. Exp Toxicol Pathol 2013;65(3):341-349.##Jha RK, Zi-rong X. Biomedical compounds from marine organisms. Mar Drugs 2004;2:123-146.##Zupo V, Jüttner F, Maibam C, Butera E, Blom J. Apoptogenic metabolites in fractions of the benthic diatom Cocconeis scutellum parva. Mar Drugs 2014;12(1):547-567.##Ramirez-gomez FJ, García-arrarás GE. Echinoderm immunity. ISJ 2010;7:211-220.##Czarkwiani A, Dylus DV, Oliveri P. Expression of skeletogenic genes during arm regeneration in the brittle star Amphiura filiformis. Gene Expr Patterns 2013;13(8):464-472.##Wang W, Hong J, Lee CO, Cho HY, Shin S, Jung JH. Bioactive metabolits from the brittle star Ophioplocus japonicus. Nat Prod Sci 2004;10(6):253-261.##Baharara J, Amini E. The potential of brittle star extracted polysaccharide in promoting apoptosis via intrinsic signaling pathway. Avicenna J Med Biotech 2015;7(4):151-158.##Chen G, Zhang P, Huang T, Yu W, Lin J, Li P, et al. Polysaccharides from Rhizopus nigricans mycelia induced apoptosis and G2/M arrest in BGC-823 cells. Carbohydr Polym 2013;97(2):800888.##Lavi I, Friesem D, Geresh S, Hadar Y, Schwartz B. An aqueous polysaccharide extract from the edible mushroom Pleurotus ostreatus induces anti-proliferative and pro-apoptotic effects on HT-29 colon cancer cells. Cancer Lett 2006;244(1):61-70.##Cao W, Li XQ, Wang X, Fan HT, Zhang XN, Hou Y, et al. A novel polysaccharide, isolated from Angelica sinensis (Oliv.) Diels induces the apoptosis of cervical cancer HeLa cells through an intrinsic apoptotic pathway. Phytomedicine 2010;17(8-9):598-605.##Pomin VH. Holothurian fucosylated chondroitin sulfate. Mar Drugs 2014;12(1988):232-254.##Gamal-Eldeen AM, Ahmed EF, Abo-Zeid MA. In vitro cancer chemopreventive properties of polysaccharide extract from the brown alga, Sargassum latifolium. Food Chem Toxicol 2009;47(6):1378-1384.##Lee K, Shin J, Nam K. Cancer chemopreventive effects of starfish polysaccharide in human breast cancer cells. Biotechnol Bioprocess Eng 2011;16(5):987-991.##Lu X, Liu W, Wu J, Li M, Wang J, Wu J, et al. A polysaccharide fraction of adlay seed ( Coix lachryma - jobi L.) induces apoptosis in human non-small cell lung cancer A549 cells. Biochem Biophys Res Commun 2013;430(2):846-851.##Wang Z, Lu C, Wu C, Xu M, Kou X, Kong D et al. Polysaccharide of Boschniakia rossica induces apoptosis on laryngeal carcinoma Hep2 cells. Gene 2014;536(1):203-206.##Thangam R, Sathuvan M, Poongodi A, Suresh V, Pazhanichamy K, Sivasubramanian S, et al. Activation of intrinsic apoptotic signaling pathway in cancer cells by Cymbopogon citratus polysaccharide fractions. Carbohydr Polym 2014;107:138-150.##Yazdanpanahi N, Behbahani M, Yektaeiana A. Effect of Boswellia thurifera gum methanol extract on cytotoxicity and p53 gene expression in human breast cancer cell line. Iran J Pharm Res 2014;13(2):719-724.##Zhang X, Yu H. Matrine inhibits diethylnitrosamine-induced HCC proliferation in rats through inducing apoptosis via p53, Bax-dependent caspase-3 activation pathway and down-regulating MLCK overexpressio. Iran J Pharm Res 2016;15(2):491-499.##

MIG1 Glucose Repression in Metabolic Processes of Saccharomyces cerevisiae: Genetics to Metabolic Engineering 2 2 Background: Although Saccharomyces cerevisiae has several industrial applications, there are still fundamental problems associated with sequential use of carbon sources. As such, glucose repression effect can direct metabolism of yeast to preferably anaerobic conditions. This leads to higher ethanol production and less efficient production of recombinant products. The general glucose repression system is constituted by MIG1, TUP1 and SSN6 factors. The role of MIG1 is known in glucose repression but the evaluation of effects on aerobic/anaerobic metabolism by deletion of MIG1 and constructing an optimal strain brand remains unclear and an objective to be explored.  Methods: To find the impact of MIG1 in induction of glucose-repression, the Mig1 disruptant strain (∆MIG1) was produced for comparing with its congenic wild-type strain (2805). The analysis approached for changes in the rate of glucose consumption, biomass yield, cell protein contents, ethanol and intermediate metabolites production. The MIG1 disruptant strain exhibited 25% glucose utilization, 12% biomass growth rate and 22% protein content over the wild type. The shift to respiratory pathway has been demonstrated by 122.86 and 40% increase of glycerol and pyruvate production, respectively as oxidative metabolites, while the reduction of fermentative metabolites such as acetate 35.48 and ethanol 24%.  Results: Results suggest that ∆MIG1 compared to the wild-type strain can significantly present less effects of glucose repression.  Conclusion: The constructed strain has more efficient growth in aerobic cultivations and it can be a potential host for biotechnological recombinant yields and industrial interests. 215 220 Iraj Alipourfard Institute of Chemical Biology, Faculty of Natural Sciences and Engineering, Ilia State UniversityCenter of Pharmaceutical Sciences, Faculty of Life Sciences, University of Vienna Institute of Chemical Biology, Faculty of Natural Sciences and Engineering, Ilia State UniversityCenter of Pharmaceutical Sciences, Faculty of Life Sciences, University of Vienna GeorgiaAustria Nelly Datukishvili Institute of Chemical Biology, Faculty of Natural Sciences and Engineering, Ilia State University Institute of Chemical Biology, Faculty of Natural Sciences and Engineering, Ilia State University Georgia Salar Bakhtiyari Department of Clinical Biochemistry, Faculty of Medicine, Ilam University of Medical Sciences Department of Clinical Biochemistry, Faculty of Medicine, Ilam University of Medical Sciences Iran Karimeh Haghani Department of Clinical Biochemistry, Faculty of Medicine, Ilam University of Medical Sciences Department of Clinical Biochemistry, Faculty of Medicine, Ilam University of Medical Sciences Iran Laura Di Renzo Section of Clinical Nutrition and Nutrigenomics, Department of Biomedicine and Prevention, University of Rome Tor Vergata Section of Clinical Nutrition and Nutrigenomics, Department of Biomedicine and Prevention, University of Rome Tor Vergata Italy Renata de Miranda Faculty of Applied Medical-Surgical Sciences, University of Rome Tor VergataCAPES Foundation, Ministry of Education of Brazil Faculty of Applied Medical-Surgical Sciences, University of Rome Tor VergataCAPES Foundation, Ministry of Education of Brazil ItalyBrazil David Mikeladze Institute of Chemical Biology, Faculty of Natural Sciences and Engineering, Ilia State University Institute of Chemical Biology, Faculty of Natural Sciences and Engineering, Ilia State University Georgia Saccharomyces cerevisiaeHomologous recombinationMetabolic pathwaysYeasts 10377.pdf Xu JR, Zhao XQ, Liu CG, Bai FW. Improving xylose utilization of Saccharomyces cerevisiae by expressing the MIG1 mutant from the self-flocculating yeast SPSC01. Protein Pept Lett 2018;25(2):202-207.##Galello F, Pautasso C, Reca S, Cañonero L, Portela P, Moreno S, et al. Transcriptional regulation of the protein kinase a subunits in Saccharomyces cerevisiae during fermentative growth. Yeast 2017;34(12):495-508.##Klein CJL, Rasmussen JJ, Rønnow B, Olsson L, Nielsen J. Investigation of the impact of MIG1 and MIG2 on the physiology of Saccharomyces cerevisiae. J Biotech 1999;68(2-3):197-212.##Nehlin JO, Ronne H. Yeast MIG1 repressor is related to the mammalian early growth response and Wilms tumour finger proteins. EMBO J 1990;9(9):2891-2898.##Dowzer CEA, Kelly LM. Analysis of the creA gene, a regulator of carbon catabolite repression in Aspergillus nidulans. Mol Cell Biol 1991;11(11):5701-5709.##Entian KD, Schuller HJ. Glucose repression (carbon catabolite repression) in yeast. In: Zimmermann FK, Entian KD (eds). Yeast Sugar Metabolism. Lancaster, Pennsylvania; Technomic Publishing Company; 1997. p. 409-434.##Shashkova S, Wollman AJM, Leake MC, Hohmann S. The yeast Mig1 transcriptional repressor is dephosphorylated by glucose-dependent and -independent mechanisms. FEMS Microbiol Lett 2017;364(14):fnx133.##Westholm OJ, Nordberg N, Murén E, Ameur A, Komorowski J, Ronne H. Combinatorial control of gene expression by the three yeast repressors Mig1, Mig2 and Mig3. BMC Genomics 2008;9:601.##Atkins D, Arndt GM, Izant JG. Antisense gene expression in yeast. Biol Chem Hoppe-Seyler 1994;375 (11):721-729. ##Olsson L, Larsen ME, Rønnow B, Mikkelsen JD, Nielsen J. Silencing MIG1 in Saccharomyces cerevisiae: Effects of antisense MIG1 expression and MIG1 gene disruption. Appl Environ Microbiol 1997;63(6):2366-2371.##Rose M, Winston F, Hieter P. Methods in yeast genetics: a laboratory course manual. Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press; 1990. 198 p.##Verduyn C, Postma E, Scheffers WA, van Dijken JP. Physiology of Saccharomyces cerevisiae in anaerobic glucose-limited chemostat cultures. J Gen Microbiol 1990;136(3):395-403.##Klein CJL, Olsson L, Rønnow B, Mikkelsen JD, Nielsen J. Alleviation of glucose repression of maltose metabolism by MIG1 disruption in Saccharomyces cerevisiae. Appl Environ Microbiol 1996;62(12):4441-4449.##Ausubel FM, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, et al. Short protocols in molecular biology, 3rd ed. New York: John Wiley & Sons;1995. 836 p.##Rothstein RJ. One step gene disruption in yeast. Methods Enzymol 1983;101:202-211.##Sambrook J, Fritsch EF, Maniatis T. Molecular cloning: a laboratory manual, 2nd ed. Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press; 1989. 1626 p.##Klein CJL, Olsson L, Nielsen J. Glucose control in Saccharomyces cerevisiae: the role of MIG1 in metabolic functions. 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Phylogenetic Analysis of Hepatitis B Virus among Household Members with HBV Chronic Infection 2 2 Background: Intrafamilial spread of Hepatitis B virus (HBV) infection in Iran has only been investigated with serological testing without using molecular studies as the most informative and definitive type of analysis.  Methods: In the present study, intrafamilial transmission of HBV among family members of Iranian index HBsAg carriers was investigated using phylogenetic analysis of the S region of the viral genome. Nested polymerase chain reaction was used for detection of HBV DNA in serum samples from 22 index and 43 contact patients with chronic HBV infection. HBV DNA was detected in 37 samples (14 indexes, 23 contacts). The S gene region of the DNA isolates was subjected to direct sequencing and phylogenetic analysis by Bioedit, Mega and Phylip programs.  Results: All isolates (from 26 patients) were clustered with genotype D, of which 24 strains were of subgenotype D1, subtype ayw2, while 2 additional strains were of subgenotype D2, subtype ayw3. Evidence of intrafamilial transmission of the virus was found in 8 families studied phylogenetically. Overall, 60 changes were detected in the amino acid sequences of the surface antigen protein in 23 patients. Four premature stop codons occurred in 3 isolates at residues 69 and 182. Seven out of 8 families displayed 25−100% common amino acid substitutions among their members.  Conclusion: Our data corroborated intrafamilial transmission of HBV, as evidenced by concordant HBV genotype among household members, viral sequence homology and close genetic relatedness of the strains on the phylogenetic tree, and horizontal transmission of S gene mutations among family members. 221 228 Shahnaz Sali Department of Infectious Diseases and Tropical Medicine, Shahid Beheshti University of Medical Sciences Department of Infectious Diseases and Tropical Medicine, Shahid Beheshti University of Medical Sciences Iran Shirin Azarmmanesh Department of Infectious Diseases and Tropical Medicine, Shahid Beheshti University of Medical Sciences Department of Infectious Diseases and Tropical Medicine, Shahid Beheshti University of Medical Sciences Iran Hediyeh Ghalikhani Department of Immunology, Faculty of Medicine, Shahid Beheshti University of Medical Sciences Department of Immunology, Faculty of Medicine, Shahid Beheshti University of Medical Sciences Iran Maryam Vaezjalali Department of Microbiology, Faculty of Medicine, Shahid Beheshti University of Medical Sciences Department of Microbiology, Faculty of Medicine, Shahid Beheshti University of Medical Sciences Iran GenotypeHepatitis B virusPhylogenetic analysisSurface antigenTransmission 10378.pdf Kew MC. Hepatitis B virus: epidemiology and clinical features of related cancer. In: Hudnall SD, editor. Viruses and human cancer. New York: Springer; 2014. p. 133-165.##Kuo A, Gish R. Chronic hepatitis B infection. Clin liver Dis 2012;16(2):347-369.##Ma L, Alla NR, Li X, Mynbaev OA, Shi Z. Mother‐to‐ child transmission of HBV: review of current clinical management and prevention strategies. Rev Med Virol 2014;26(4):396-406.##Ucmak H, Faruk Kokoglu O, Celik M, Ergun UG. Intra-familial spread of hepatitis B virus infection in eastern Turkey. Epidemiol Infect 2007;135(8):1338-1343.##Alavian SM, Taheri S. A Global Perspective on the Intrafamilial Transmission of Hepatitis B Virus Infection. Int J Travel Med Glob Health 2012;1(1):22-26.##Hatami H, Salehi M, Sanei E, Khosravi S, Alavian SM. Intra-familial transmission of hepatitis B virus infection in Zahedan. Iran Red Crescent Med J 2013;15(1):4-8.##Mansour-Ghanaei F, Joukar F, Yaseri M, Soati F, Atrkar-Roushan Z. Intrafamilial spread of hepatitis B virus in Guilan Province-North of Iran. Int J Mol Epidemiol Genet 2013;4(4):250-257.##Dumpis U, Holmes EC, Mendy M, Hill A, Thursz M, Hall A, et al. Transmission of hepatitis B virus infection in Gambian families revealed by phylogenetic analysis. J Hepatol 2001;35(1):99-104.##Lin CL, Kao JH, Chen BF, Chen PJ, Lai MY, Chen DS. Application of hepatitis B virus genotyping and phylogenetic analysis in intrafamilial transmission of hepatitis B virus. Clin Infect Dis 2005;41(11):1576-1581.##Alavian SM, Hajarizadeh B, Ahmadzad-Asl M, Kabir A, Bagheri-Lankarani K. Hepatitis B virus infection in Iran: a systematic review. Hepat Mon 2008;8(4):281-294.##Toukan AU. Hepatitis B in the middle east: aspects of epidemiology and liver disease after infection. Gut 1996; 38 Suppl 2:S2-S4.##Sofian M, Banifazl M, Ziai M, Aghakhani A, Farazi AA, Ramezani A. Intra-familial transmission of hepatitis B virus infection in Arak, central Iran. 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Surface gene mutations of hepatitis B virus among high-risk patients with occult hepatitis B virus infection. Diagn Microbiol Infect Dis 2010;66(3):285-291.##Asli M, Kandelouei T, Rahimyan K, Davoodbeglou F, Vaezjalali M. Characterization of occult hepatitis B infection among injecting drug users in Tehran, Iran. Hepat Mon 2016;16(3):e34763.##Kim BK, Revill PA, Ahn SH. HBV genotypes: relevance to natural history, pathogenesis and treatment of chronic hepatitis B. Antivir Ther 2011;16(8):1169-1186.##Norder H, Couroucé AM, Coursaget P, Echevarria JM, Lee SD, Mushahwar IK, et al. Genetic diversity of hepatitis B virus strains derived worldwide: genotypes, subgenotypes, and HBsAg subtypes. Intervirology 2004;47(6):289-309.##Pourkarim MR, Amini-Bavil-Olyaee S, Kurbanov F, Van Ranst M, Tacke F. Molecular identification of hepatitis B virus genotypes/subgenotypes: Revised classification hurdles and updated resolutions. World J Gastroenterol 2014;20(23):7152-7168.##Moradi A, Zhand S, Ghaemi A, Javid N, Tabarraei A. Mutations in the S gene region of hepatitis B virus genotype D in Golestan province-Iran. Virus Genes 2012;44(3):382-387.##Lin CL, Kao JH. The clinical implications of hepatitis B virus genotype: recent advances. J Gastroenterol Hepatol 2011;26 Suppl 1:123-130.##Lin CL, Kao JH. Intrafamilial transmission of hepatitis B virus infection. J Gastroenterol Hepatol 2007;22(5):765-766.##Lin HJ, Lai CL, Lau JY, Chung HT, Lauder IJ, Fong MW. Evidence for intrafamilial transmission of hepatitis B virus from sequence analysis of mutant HBV DNAs in two Chinese families. Lancet 1990;336(8709):208-212.##Datta S, Banerjee A, Chandra PK, Chowdhury A, Chakravarty R. Genotype, phylogenetic analysis, and transmission pattern of occult hepatitis B virus (HBV) infection in families of asymptomatic HBsAg carriers. 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Effect of Recombinant Helicobacter Outer Membrane Protein H (HopH) on Nitric Oxide Production by Peripheral Macrophage in BALB/c Mice 2 2 Background: Some products of bacteria are reported as an immunomodulator. The Helicobacter pylori (H. pylori) outer membrane proteins play an important role in stimulation of immune system. The present study was performed to determine the in vitro effect of recombinant HopH of H. pylori on Nitric Oxide (NO) production and viability of mouse peritoneal macrophages. Methods: H. pylori recombinant HopH was produced in this study. Mice peritoneal macrophages were purified and cultured. Different concentrations of recombinant HopH were used for stimulation of macrophages in order to evaluate NO production. The cell viability was detected by MTT assay. NO amounts released in to the supernatants of cultured macrophages and LPS-stimulated macrophages (10 μg/ml) were detected by Griess reagent. Results: Results demonstrated that the suppressive effect of high concentrations of recombinant HopH on NO release and the stimulation effect of protein was shown in 15 µg/ml, compared to the control group. NO stimulation was significant in all the concentrations of LPS stimulated with HopH groups. Conclusion: According to our findings, recombinant HopH has a toxic effect in high concentration on cell. So it can be an anticancer candidate. 229 233 Masoumeh Navidinia Faculty of Allied Medical Sciences, Shahid Beheshti University of Medical Sciences Faculty of Allied Medical Sciences, Shahid Beheshti University of Medical Sciences Iran Neda Soleimani Departments of Microbiology and Microbial Biotechnology and Biotechnology, Faculty of Life Sciences and Biotechnology, Shahid Beheshti University Departments of Microbiology and Microbial Biotechnology and Biotechnology, Faculty of Life Sciences and Biotechnology, Shahid Beheshti University Iran Narges Bodagh abadi Department of Genetics, Faculty of Biological Sciences, Tarbiat Modares University Department of Genetics, Faculty of Biological Sciences, Tarbiat Modares University Iran Helicobacter pyloriMacrophageRecombinant HopH 10379.pdf Marin ML, Tejada-Simon MV, Lee J, Ustunol Z, Pestka JJ. Effects of Lactobacillus spp. on cytokine production by RAW 264.7 macrophages and EL-4 thymoma cell lines. J Food Prot 1997;60(11):1364-1370.##Ribeiro-Dias F, Russo M, Nascimento FRF, Barbuto JAM, Timenetsky J, Jancar S. Thioglycollate-elicited murine macrophages are cytotoxic to Mycoplasma arginine infected YAC-1 tumor cells. Braz J Med Biol Res 1998;31(11):1425-1428.##Michetti P, Kreiss C, Kotloff KL, Porta N, Blanco JC, Bachmann D, et al. Oral immunization with urease and Escherichia coli heat-labile enterotoxin is safe and immunogenic in Helicobacter pylori-infected adults. Gastroenterology 1999;116(4):804-812. ##Zhang XQ, Lin SR. Progress in research on the relationship between Hp and stomach cancer. World Chinese J Digestology 2000;8:206-207.##Meyer JM, Silliman NP, Dixon CA, Siepman NY, Sugg JE, Hopkins RJ. Helicobacter pylori and early duodenal ulcer status post-treatment: a review. Helicobacter 2001; 6(2):84-92. ##Watanabe T, Tada M, Nagai H, Sasaki S, Nakao M. Helicobacter pylori infection induces gastric cancer in Mongolian Gerbils. Gastroenterol 1998;115(3):642-648. ##Honda S, Fujioka T, Tokieda M, Satoh R, Nishizono A, Nasu M. Development of Helicobacter pylori-induced gastric carcinoma in Mongolian Gerbils. Cancer Res 1998;58(19):4255-4259. ##Suerbaum S, Michetti P. Helicobacter pylori infection. N Engl J Med 2002;347(15):1175-1186.##Yamaoka Y, Kikuchi S, el-Zimaity HM, Gutierrez O, Osato MS, Graham DY. Importance of Helicobacter pylori oipA in clinical presentation, gastric inflammation, and mucosal interleukin 8 production. Gastroenterology 2002;123(2):414-424. ##Cover TL. Role of Helicobacter pylori outer membrane proteins in gastroduodenal disease. J Infect Dis 2006;194:1343-1345.##Dossumbekova A, Prinz C, Mages J, Lang R, Kusters JG, Van Vliet AH, Reindl W, et al. Helicobacter pylori HopH (OipA) and bacterial pathogenicity: genetic and functional genomic analysis of hopH gene polymorphisms. J Infect Dis 2006;194(10):1346-1355. ##Liu J, He C, Chen M, Wang Z, Xing C, Yuan Y. Association of presence/absence and on/off patterns of Helicobacter pylori oipA gene with peptic ulcer disease and gastric cancer risks: a meta-analysis. BMC Infect Dis 2013;13(1):555. ##Hajimoradi M, Daneshmandi S, Mohammad Hassan Z, Roudbary M. Effect of shark liver oil on peritoneal murine macrophages in responses to killed-Candida albicans. IJBMS 2009;12(3-4):179-183.##Daneshmandi S, Hajimoradi M, Soleimani N, Sattari M. Modulatory effect of Acetobacter xylinum cellulose on peritoneal macrophages. Immunopharmacol Immuno-toxicol 2011;33(1):164-168. ##Soleimani N, Daneshmandi S, Sattari M, Pourfathollah AA. Immuno-modulatory and anti-tumor effects of Cuminum cyminum essential oil. Arak Med Univ J 2011;13(4):22-29.##Amedei A, Cappon A, Codolo G, Cabrelle A, Polenghi A, Benagiano M, et al. The neutrophil-activating protein of Helicobacter pylori promotes Th1 immune responses. J Clin Invest 2006;116(4):1092-1101. ##D'Elios MM, Amedei A, Cappon A, Del Prete G, de Bernard M. The neutrophil-activating protein of Helicobacter pylori (HP-NAP) as an immune modulating agent. FEMS Immunol Med Microbiol 2007;50(2):157-164. ##Teymournejad O, Mobarez AM, Hassan ZM, Moazzeni SM, Yakhchali B, Eskandari V. In silico prediction of exposure amino acid sequences of outer inflammatory protein A of Helicobacter pylori for surface display on Eschierchia coli. Indian J Hum Genet 2012;18(1):83-86. ##Soleimani N, Mohabati Mobarez A, Tavakoli-Yaraki M, Farhangi B. Evaluation of nitric oxide production and proliferation activity of recombinant Bacterioferritin of Helicobacter pylori on macrophages. Microb Pathog 2016;100:149-153. ##

Effect of Hydroalcoholic Ginger Extract on Brain HMG-CoA Reductase and CYP46A1 Levels in Streptozotocin-induced Diabetic Rats 2 2 Background: Patients with diabetes present with lipid disorders, including hypercholesterolemia, which can be a high-risk factor for atherosclerosis. Recently, increasing interest has been focused on anti-lipidemic function of herbal medicines, especially Zingiber officinale (known as ginger), in diabetes. However, the mechanism underlying the effect of ginger on some players involved in cholesterol homeostasis of Central Nervous System (CNS) among diabetic patients remains unclear. To our knowledge, this is the first study to investigate the effect of ginger on brain regulation of Hydroxymethylglutaryl-CoA Reductase (HMG-CoA reductase) and Cholesterol 24-hydroxylase (CYP46A1), which provides a rational model for understanding brain dyslipidemia mechanisms associated with diabetes.  Methods: Brains of rats were isolated from four groups: control, non-treated diabetic, and treated diabetic groups receiving 200 or 400 mg/kg of hydroalcoholic extracts of ginger for eight weeks. HMG-CoA reductase and CYP46A1 levels in brain homogenates were determined by western-blot technique.  Results: Ginger root extract caused a significant decrease in HMG-CoA reductase and an increase in CYP46A1 levels in treated diabetic groups compared to diabetic control. In comparison to diabetic group, these effects were more remarkable with 400 mg/kg concentration of ginger extract.  Conclusion: The findings showed that ginger extract has a regulatory effect on proteins involved in cholesterol homeostasis in CNS by a significant down- and up-regulation of HMG-CoA reductase and CYP46A1 levels, respectively. It can be suggested that adding ginger to daily diet of diabetic patients has useful effects and may ameliorate diabetes complications. 234 238 Shirin Azizidoost Department of Biochemistry, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences Department of Biochemistry, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences Iran Zahra Nazeri Department of Biochemistry, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences Department of Biochemistry, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences Iran Asma Mohammadi Department of Biochemistry, Abadan University of Medical Sciences Department of Biochemistry, Abadan University of Medical Sciences Iran Ghorban Mohammadzadeh Hyperlipidemia Research Center, Department of Clinical Biochemistry, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences Hyperlipidemia Research Center, Department of Clinical Biochemistry, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences Iran Maryam Cheraghzadeh Department of Biochemistry, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences Department of Biochemistry, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences Iran Alireza Jafari Department of Biochemistry, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences Department of Biochemistry, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences Iran Alireza Kheirollah Department of Biochemistry, Cellular and Molecular Research Center, Ahvaz Jundishapur University of Medical Sciences Department of Biochemistry, Cellular and Molecular Research Center, Ahvaz Jundishapur University of Medical Sciences Iran BrainCholesterol 24-hydroxylaseDiabetes mellitusGingerHydroxymethylglutaryl-CoA reductases 10380.pdf Mustafa SS, Eid NI, Jafri S, El-Latif HAA, Ahmed HM. Insulinotropic effect of aqueous ginger extract and aqueous garlic extract on the isolated perfused pancreas of streptozotocin induced diabetic rats. Pakistan J Zool 2007;39(5):279-284.##Al-Azhary DB. Ginger enhances antioxidant activity and attenuates atherogenesis in diabetic cholesterol-fed rats. Aust J Basic Appl Sci 2011;5(12):2150-2158.##Luchsinger JA, Tang MX, Stern Y, Shea S, Mayeux R. Diabetes mellitus and risk of Alzheimer's disease and dementia with stroke in a multiethnic cohort. Am J Epidemiol 2001;154(7):635-641.##Martins I, Hone E, Foster J, Sünram-Lea S, Gnjec A, Fuller S, et al. Apolipoprotein E, cholesterol metabolism, diabetes, and the convergence of risk factors for Alzheimer's disease and cardiovascular disease. Mol Psychiatry 2006;11(8):721-736.##Bruce DG, Casey GP, Grange V, Clarnette RC, Almeida OP, Foster JK, et al. Cognitive impairment, physical disability and depressive symptoms in older diabetic patients: the Fremantle Cognition in Diabetes Study. Diabetes Res Clin Pract 2003;61(1):59-67.##Stewart R, Liolitsa D. Type 2 diabetes mellitus, cognitive impairment and dementia. Diabet Med 1999;16(2):93-112.##Pihlajamäki J, Gylling H, Miettinen TA, Laakso M. Insulin resistance is associated with increased cholesterol synthesis and decreased cholesterol absorption in normoglycemic men. J Lipid Res 2004;45(3):507-512.##Simonen PP, Gylling HK, Miettinen TA. Diabetes contributes to cholesterol metabolism regardless of obesity. Diabet Care 2002;25(9):1511-1515.##Suzuki R, Lee K, Jing E, Biddinger SB, McDonald JG, Montine TJ, et al. Diabetes and insulin in regulation of brain cholesterol metabolism. Cell Metab 2010;12(6):567-579.##Dietschy JM, Turley SD. Thematic review series: brain Lipids. Cholesterol metabolism in the central nervous system during early development and in the mature animal. J Lipid Res 2004;45(8):1375-1397.##Sun H, Yuan Y, Sun ZL. Cholesterol contributes to diabetic nephropathy through SCAP-SREBP-2 pathway. Int J Endocrinol 2013;2013.##Wang Z, Jiang T, Li J, Proctor G, McManaman JL, Lucia S, et al. Regulation of renal lipid metabolism, lipid accumulation, and glomerulosclerosis in FVBdb/db mice with type 2 diabetes. Diabetes 2005;54(8):2328-2335.##Yuan Y, Zhao L, Chen Y, Moorhead JF, Varghese Z, Powis SH, et al. Advanced glycation end products (AGEs) increase human mesangial foam cell formation by increasing Golgi SCAP glycosylation in vitro. Am J Physiol Renal Physiol 2011;301(1):F236-F43.##Brunham LR, Kruit JK, Verchere CB, Hayden MR. Cholesterol in islet dysfunction and type 2 diabetes. J Clin Invest 2008;118(2):403-408.##Brunham LR, Kruit JK, Pape TD, Timmins JM, Reuwer AQ, Vasanji Z, et al. β-cell ABCA1 influences insulin secretion, glucose homeostasis and response to thiazolidinedione treatment. Nat Med 2007;13(3):340-347. ##Li Y, Tran VH, Duke CC, Roufogalis BD. Preventive and protective properties of Zingiber officinale (ginger) in diabetes mellitus, diabetic complications, and associated lipid and other metabolic disorders: a brief review. Evid Based Complement Alternat Med 2012;2012:516870.##Ojewole JA. Analgesic, antiinflammatory and hypoglycaemic effects of ethanol extract of Zingiber officinale (Roscoe) rhizomes (Zingiberaceae) in mice and rats. Phytother Res 2006;20(9):764-772.##Singh Rajput DP, Shah JY, Singh P, Jain S. Evaluation of dyslipidemia in type 2 diabetes mellitus. Asian J Med Sci 2015;6(6):16-19.##Robert J, Cheng WH, Hayat A, Ward-Able T, Wellington CL. High-density lipoproteins at the interface between central nervous system and plasma lipoprotein metabolism. Clin Lipidol 2017;10(1):69-81.##Djelti F, Braudeau J, Hudry E, Dhenain M, Varin J, Bieche I, et al. CYP46A1 inhibition, brain cholesterol accumulation and neurodegeneration pave the way for Alzheimer’s disease. Brain 2015;138(8):2383-2398.##Boussicault L, Alves S, Lamazière A, Planques A, Heck N, Moumné L, et al. CYP46A1, the rate-limiting enzyme for cholesterol degradation, is neuroprotective in Huntington’s disease. Brain. 2016;139(3):953-970.##DeBose-Boyd RA. Feedback regulation of cholesterol synthesis: sterol-accelerated ubiquitination and degradation of HMG CoA reductase. Cell Res 2008;18(6):609-621.##Wang Y, Muneton S, Sjövall J, Jovanovic JN, Griffiths WJ. The effect of 24S-hydroxycholesterol on cholesterol homeostasis in neurons: quantitative changes to the cortical neuron proteome. J Proteome Res 2008;7(4):1606-1614.##Elshater A, Salman MM, Moussa MM. Effect of ginger extract consumption on levels of blood glucose, lipid profile and kidney functions in alloxan induced-diabetic rats. Egypt Acad J Biolog Sci 2009;2(1):153-162.##El-Akabawy G, El-Kholy W. Neuroprotective effect of ginger in the brain of streptozotocin-induced diabetic rats. Ann Anat 2014;196(2-3):119-128.##

Investigation of the Association between 5-Hydroxytryptamine Transporter Gene-Linked Polymorphic Region with Type 2 Diabetes Mellitus, Obesity and Biochemical Profiles of Serum in Iranian Population 2 2 Background: Type 2 Diabetes Mellitus (T2DM) is a serious problem in the world. 5-Hydroxytryptamine (5-HT, serotonin) plays an important role in obesity, glucose control and insulin resistance. The polymorphism of the serotonin transporter gene linked promoter region (5-HTTLPR) might influence 5-HTT expression and serotonin uptake. The polymorphism results in two alleles of L (Long) and S (Short). The aim of the present study was to evaluate the association between 5-HTTLPR genotypes in type 2 diabetes mellitus (T2DM), obesity as well as serum biochemical profiles in Iranian population from 2012 until 2015. Methods: 180 patients with T2DM and 180 controls were selected and the frequency of S and L alleles was determined by PCR. Then, the relationship between genotypes, body mass index (BMI) and serum biochemical variables was investigated. Results: The frequency of S and L alleles in experimental and control groups was the same [for the L allele p=0.754, OR (95%CI)=1.103 (0.597 to 2.041) and for the S allele p=0.906, OR (95%CI)=(0.490 to 1.676)]. However, the mean triglyceride, cholesterol, LDL-C, systolic and diastolic blood pressure levels in the diabetic subjects with LL genotype were significantly higher than LS and SS genotypes (p<0.001) in this population. Conclusion: The L allele of 5-HTTLPR was related to the increased serum lipids and blood pressure in the diabetic patients. However, there was no relationship between the polymorphism of 5-HTTLPR L/S and T2DM in Iranian population.  239 244 Azizeh Asadzadeh Department of Biology, University of Nourdanesh Institute of Higher Education, Meymeh Department of Biology, University of Nourdanesh Institute of Higher Education, Meymeh Iran Hooria Seyedhosseini Ghaheh Young Researchers and Elites Club, Science and Research Branch, Islamic Azad University Young Researchers and Elites Club, Science and Research Branch, Islamic Azad University Iran Fatemeh Sholehvar Department of Biology, Faculty of Science, Zand Institute of Higher Education Department of Biology, Faculty of Science, Zand Institute of Higher Education Iran Mohammadali Takhshid Diagnostic Laboratory Sciences and Technology Research Center, School of Paramedical Sciences, Shiraz University of Medical Sciences Diagnostic Laboratory Sciences and Technology Research Center, School of Paramedical Sciences, Shiraz University of Medical Sciences Iran Mohammad Mehdi Naghizadeh Noncommunicable Diseases Research Center, Fasa University of Medical Sciences Noncommunicable Diseases Research Center, Fasa University of Medical Sciences Iran 5-HTT gene5-HTTLPR polymorphismSerotoninType 2 diabetes mellitus 10381.pdf Lau T, Schloss P. Differential regulation of serotonin transporter cell surface expression. Wiley Interdisciplinary Reviews: Membrane Transport and Signaling 2012;1:259.##Jenkins TA, Nguyen JC, Polglaze KE, Bertrand PP. Influence of tryptophan and serotonin on mood and cognition with a possible role of the gut-brain axis. Nutrients 2016;8(1).pii:E56.##Lesch KP, Bengel D, Heils A, Sabol SZ, Greenberg BD, Petri S, et al. Association of anxiety-related traits with a polymorphism in the serotonin transporter gene regulatory region. Science 1996;274(5292):1527-1531.##Ni W, Watts SW. 5-Hydroxytryptamine in cardiovascular system: Focus on the serotonin transporter (SERT). Clin Exp Pharmacol Physiol 2006;33(7):575-583. ##Sookoian S, Gianotti TF, Gemma C, Burgueño A, Pirola CJ. Contribution of the functional 5-HTTLPR variant of the SLC6A4 gene to obesity risk in male adults. Obesity 2008;16(2):488-491.##Iordanidou M, Tavridou A, Petridis I, Arvanitidis KI, Christakidis D, Vargemezis V, et al. The serotonin transporter promoter polymorphism (5-HTTLPR) is associated with type 2 diabetes. Clinica Chimica Acta 2010;411(3-4):167-171.##Lam DD, Heisler LK. Serotonin and energy balance: molecular mechanisms and implications for type 2 diabetes. Expert Rev Mol Med 2007;9(5):1-24.##Zhou L, Sutton GM, Rochford JJ, Semple RK, Lam DD, Oksanen LJ, et al. Serotonin 2C receptor agonists improve type 2 diabetes via melanocortin-4 receptor signaling pathways. Cell Metab 2007;6(5):398-405.##Kyzar EJ, Stewart AM, Kalueff AV. Effects of LSD on grooming behavior in serotonin transporter heterozygous (Sert+/−) mice. Behav Brain Res 2016;296:47-52.##Smeraldi E, Serretti A, Artioli P, Lorenzi C, Catalano M. Serotonin transporter gene-linked polymorphic region: possible pharmacogenetic implications of rare variants. Psychiatr Genet 2006;16(4):153-158.##Avula R, Rand A, Black J, O'Kane D. Simultaneous genotyping of multiple polymorphisms in human serotonin transporter gene and detection of novel allelic variants. Transl Psychiatry 2011;1(8):e32.##Lesch K-P, Balling U, Gross J, Strauss K, Wolozin B, Murphy D, et al. Organization of the human serotonin transporter gene. J Neural Transm Gen Sect 1994;95(2):157-162.##Heils A, Teufel A, Petri S, Stöber G, Riederer P, Bengel D, et al. Allelic variation of human serotonin transporter gene expression. J Neurochem 1996;66(6):2621-2624.##Mortensen OV, Thomassen M, Larsen MB, Whittemore SR, Wiborg O. Functional analysis of a novel human serotonin transporter gene promoter in immortalized raphe cells. Brain Res Mol Brain Res 1999;68(1-2):141-148.##Bonasera SJ, Tecott LH. Mouse models of serotonin receptor function: toward a genetic dissection of serotonin systems. Pharmacol Ther 2000;88(2):133-142.##Nonogaki K, Strack AM, Dallman MF, Tecott LH. Leptin-independent hyperphagia and type 2 diabetes in mice with a mutated serotonin 5-HT2C receptor gene. Nat Med 1998;4(10):1152-1156.##Peralta-Leal V, Leal-Ugarte E, Meza-Espinoza JP, Dávalos-Rodríguez IP, Bocanegra-Alonso A, Acosta-González RI, et al. Association of a serotonin transporter gene (SLC6A4) 5-HTTLPR polymorphism with body mass index categories but not type 2 diabetes mellitus in Mexicans. Genet Mol Biol 2012;35(3):589-593.##Hameed A, Ajmal M, Nasir M, Ismail M. Genetic association analysis of serotonin transporter polymorphism (5-HTTLPR) with type 2 diabetes patients of Pakistani population. Diabetes Res Clin Pract 2015;108(1):67-71.##Nazem H, Takhshid MA, Tabei SMB, Sholevar F, Entezam M, Manoochehri J. [Investigation of association between serotonin transporter gene and type 2 diabetes mellitus]. Iran J Diabetes Lipid Disord 2010;1:84-89. Persian.##Sookoian S, Gemma C, García SI, Gianotti TF, Dieuzeide G, Roussos A, et al. 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Sericin Ameliorates the Capacitation State and Chromatin Integrity of Frozen-Thawed Stallion Spermatozoa by Reducing Oxidative Stress 2 2 Background: In the process of sperm cryopreservation, apart from cryoinjury, the production of Reactive Oxygen Species (ROS) can adversely affect the integrity of chromatin and cellular membranes. Addition of natural antioxidants to freezing me-dium is an approach to reduce the destructive effects of ROS on sperm.  Methods: In this study, during 60 min of cooling process, the ejaculates of five stallions were diluted in the following media: INRA 82 medium as Control (C), INRA 82 medium supplemented with 0.25% Sericin (S), INRA 82 medium supplemented with 1.5 mM Glutathione (G), and INRA 82 medium supplemented with 0.25% Sericin+1.5 mM Glutathione (S+G). Results: In the frozen/thawed sericin supplemented group, while the integrity of DNA and the activity of catalase and Glutathione Peroxidase (GPx) were increased, the lipid peroxidation and midpieceab normality decreased, compared with other groups (p<0.05). The proportions of sperms with abnormal head in group S and the sperm with distal droplet in G and S+G groups decreased, compared with group C (p<0.05). In CTC assay, the percentage of capacitated spermatozoa in treatment groups was lower than control (p<0.01).  Conclusion: In conclusion, the presence of sericin in freezing medium of stallion semen could improve sperm DNA integrity and its resistance to ROS and lipid peroxidation. 245 252 Mahboobeh Heidari Nasirabadi Department of Clinical Sciences, Faculty of Veterinary Medicine, Shahrekord UniversityResearch Institute of Animal Embryo Technology, Shahrekord University Department of Clinical Sciences, Faculty of Veterinary Medicine, Shahrekord UniversityResearch Institute of Animal Embryo Technology, Shahrekord University IranIran Abolfazl Shirazi Department of Clinical Sciences, Faculty of Veterinary Medicine, Shahrekord UniversityResearch Institute of Animal Embryo Technology, Shahrekord UniversityReproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Department of Clinical Sciences, Faculty of Veterinary Medicine, Shahrekord UniversityResearch Institute of Animal Embryo Technology, Shahrekord UniversityReproductive Biotechnology Research Center, Avicenna Research Institute, ACECR IranIranIran Ali Kadivar Department of Clinical Sciences, Faculty of Veterinary Medicine, Shahrekord UniversityResearch Institute of Animal Embryo Technology, Shahrekord University Department of Clinical Sciences, Faculty of Veterinary Medicine, Shahrekord UniversityResearch Institute of Animal Embryo Technology, Shahrekord University IranIran Naser Shams-Esfandabadi Department of Clinical Sciences, Faculty of Veterinary Medicine, Shahrekord University Department of Clinical Sciences, Faculty of Veterinary Medicine, Shahrekord University Iran Abdolnaser Mohebbi Department of Clinical Sciences, Faculty of Veterinary Medicine, Shahrekord University Department of Clinical Sciences, Faculty of Veterinary Medicine, Shahrekord University Iran Ebrahim Ahmadi Research Institute of Animal Embryo Technology, Shahrekord University Research Institute of Animal Embryo Technology, Shahrekord University Iran AntioxidantsGlutathioneSericinsSpermatozoa 10382.pdf Curry MR. Cryopreservation of semen from domestic livestock. Rev Reprod 2000;5(1):46-52.##Ball BA, Vo AT, Baumber J. Generation of reactive oxygen species by equine spermatozoa. Am J Vet Res 2001;62(4):508-515.##Ball BA. Oxidative stress, osmotic stress and apoptosis: impacts on sperm function and preservation in the horse. Anim Reprod Sci 2008;107(3-4):257-267.##Baumber J, Ball BA. Determination of glutathione peroxidase and superoxide dismutase-like activities in equine spermatozoa, seminal plasma, and reproductive tissues. Am J Vet Res 2005;66(8):1415-1419.##Ball BA, Gravance CG, Medina V, Baumber J, Liu IK. Catalase activity in equine semen. Am J Vet Res 2000;61(9):1026-1030.##Kefer JC, Agarwal A, Sabanegh E. Role of antioxidants in the treatment of male infertility. Int J Urol 2009;16:449-457.##Alvarez JG, Storey BT. Taurine, hypotaurine, epinephrine and albumin inhibit lipid peroxidation in rabbit spermatozoa and protect against loss of motility. Biol Reprod 1983;29(3):548-555.##Thuwanut P, Chatdarong K, Techakumphu M, Axner E. The effect of antioxidants on motility, viability, acrosome integrity and DNA integrity of frozen-thawed epididymal cat spermatozoa. Theriogenology 2008;70(2):233-240.##Michael A, Alexopoulos C, Pontiki E, Hadjipavlou-Litina D, Saratsis P, Boscos C. Effect of antioxidant supplementation on semen quality and reactive oxygen species of frozen-thawed canine spermatozoa. Therio-genology 2007;68(2):204-212.##Baghshahi H, Riasi A, Mahdavi A, Shirazi A. Antioxidant effects of clove bud (Syzygium aromaticum) extract used with different extenders on ram spermatozoa during cryopreservation. Cryobiology 2014;69(3):482-487.##Bucak MN, Sarıözkan S, Tuncer PB, Ulutaş PA, Akçadağ Hİ. Effect of antioxidants on microscopic semen parameters, lipid peroxidation and antioxidant activities in Angora goat semen following cryopreser-vation. Small Ruminant Res 2009;81:90-95.##Foote RH, Brockett CC, Kaproth MT. Motility and fertility of bull sperm in whole milk extender containing antioxidants. Anim Reprod Sci 2002;71(1-2):13-23.##Reddy NSS, Mohanarao GJ, Atreja S. Effects of adding taurine and trehalose to a tris-based egg yolk extender on buffalo (Bubalus bubalis) sperm quality following cryopreservation. Anim Reprod Sci 2010;119(3-4):183-190.##Baumber J, Ball BA, Linfor JJ. Assessment of the cryopreservation of equine spermatozoa in the presence of enzyme scavengers and antioxidants. Am J Vet Res 2005;66:772-779. ##Li TK. The glutathione and thiol content of mammalian spermatozoa and seminal plasma. Biol Reprod 1975;12:641-646.##Bilodeau JF, Blanchette S, Gagnon C, Sirard MA. Thiols prevent H2O2-mediated loss of sperm motility in cryopreserved bull semen. Theriogenology 2001;56(2):275-286. ##Gadea J, Molla M, Selles E, Marco M, Garcia-Vazquez F, Gardon J. 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Whole Exome Sequencing of an X-linked Thrombocytopenia Patient with Normal Sized Platelets 2 2 Wiskott-Aldrich Syndrome (WAS) is a rare X-linked recessive Primary Immunodeficiency (PID) caused by mutations in WAS gene which encodes a protein known as WASp. WASp plays important roles in cytoskeletal functions that compromise multiple aspects of normal cellular activity including proliferation, phagocytosis, immune synapse formation, adhesion and directed migration. WASp defect particularly causes platelets abnormality which is presented in forms of decrease of Mean Platelet Volume (MPV) and thrombocytopenia in most WAS conditions; nevertheless, some studies reported WAS patients with a normal or large size of platelets in recent years. This phenomenon is unique and the exact mechanism of thrombocytopenia with a normal or large size of platelets is still unknown. In this study, Next Generation Sequencing (NGS) was utilized to discover the causing mutation in WAS gene; furthermore, an attempt was made to evaluate the possibility of other mutations or genes especially WASp interacting proteins and inherited platelet disorder genes in patient clinical symptoms for the purpose of understanding the origin of such unique symptom and to perform further analysis if it is required. Therefore, clinical manifestations and immunologic functions of the patient were checked and Whole Exome Sequencing (WES) was performed to analyze all exonic variations which can be associated with patient phenotypes. Finally, a novel de novo mutation in WAS gene which truncates WASp to half of its normal size was determined as the only cause of clinical manifestation. 253 258 Majid Fathi Department of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences Department of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences Iran Hojat Shahraki Department of Laboratory Sciences, Faculty of Allied Medicine, Zahedan University of Medical SciencesClinical Immunology Research Center, Zahedan University of Medical Sciences Department of Laboratory Sciences, Faculty of Allied Medicine, Zahedan University of Medical SciencesClinical Immunology Research Center, Zahedan University of Medical Sciences IranIran Edris Sharif Rahmani Department of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences Department of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences Iran Hamzeh Rahimi Department of Molecular Medicine, Biotechnology Research Center, Pasteur Institute of Iran Department of Molecular Medicine, Biotechnology Research Center, Pasteur Institute of Iran Iran Pouria omidi Department of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences Department of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences Iran Saeedeh Darvishi Department of Clinical Pathology, Faculty of Veterinary Medicine, University of Tehran Department of Clinical Pathology, Faculty of Veterinary Medicine, University of Tehran Iran Mohammad Foad Abazari Department of Genetics, Tehran Medical Sciences Branch, Islamic Azad University Department of Genetics, Tehran Medical Sciences Branch, Islamic Azad University Iran Arshad Hosseini Department of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences Department of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences Iran Blood plateletsThrombocytopeniaWhole exome sequencingWiskott-Aldrich syndrome 10383.pdf Symons M, Derry JM, Karlak B, Jiang S, Lemahieu V, McCormick F, et al. Wiskott-Aldrich syndrome protein, a novel effector for the GTPase CDC42Hs, is implicated in actin polymerization. Cell 1996;84(5):723-734.##Snapper SB, Rosen FS. The Wiskott-Aldrich syndrome protein (WASP): roles in signaling and cytoskeletal organization. Annu Rev Immunol 1999;17(1):905-929.##Thrasher AJ, Burns SO. WASP: a key immunological multitasker. Nat Rev Immunol 2010;10(3):182-192.##Ip W, Burns S, Thrasher A. WAS (Wiskott-Aldrich syndrome). 2013. http://atlasgeneticsoncology.org/Gen-es/WASID42801chXp11.html##Perry GS, Spector BD, Schuman LM, Mandel JS, Anderson VE, McHugh RB, et al. The Wiskott-Aldrich syndrome in the United States and Canada (1892-1979). J Pediatr 1980;97(1):72-78.##Candotti F. Clinical Manifestations and Pathophysiolo-gical Mechanisms of the Wiskott-Aldrich Syndrome. J Clin Immunol 2017;38(1):1-15.##Thrasher AJ. New insights into the biology of Wiskott-Aldrich syndrome (WAS). Hematology Am Soc Hematol Educ Program 2009;132-138.##Albert MH, Bittner TC, Nonoyama S, Notarangelo LD, Burns S, Imai K, et al. X-linked thrombocytopenia (XLT) due to WAS mutations: clinical characteristics, long-term outcome, and treatment options. Blood 2010; 115(16):3231-3238.##Kobayashi M, Yokoyama K, Shimizu E, Yusa N, Ito M, Yamaguchi R, et al. Phenotype-based gene analysis allowed successful diagnosis of X-linked neutropenia associated with a novel WASp mutation. Ann Hematol 2017;97(2):1-3.##Zhu Q, Zhang M, Blaese R, Derry J, Junker A, Francke U, et al. The Wiskott-Aldrich syndrome and X-linked congenital thrombocytopenia are caused by mutations of the same gene. Blood 1995;86(10):3797-3804.##Skoric D, Dimitrijevic A, Cuturilo G, Ivanovski P. Wiskott-Aldrich syndrome with macrothrombocytopenia. Indian Pediatr 2014;51(12):1015-1016.##Mazumdar J, Kanjilal S, Das A. Wiskott-Aldrich syndrome with normal-sized platelets in an eighteen-month-old boy: A rare mutation. J Pediatr Rev 2015;3(2):38-41.##Medina SS, Siqueira LH, Colella MP, Yamaguti-Hayakawa GG, Lino Duarte BK, Dos Santos Vilela MM, et al. Intermittent low platelet counts hampering diagnosis of X-linked thrombocytopenia in children: report of two unrelated cases and a novel mutation in the gene coding for the Wiskott-Aldrich syndrome protein. BMC Pediatr 2017;17(1):151.##Rocca B, Bellacosa A, De Cristofaro R, Neri G, Della Ventura M, Maggiano N, et al. Wiskott-Aldrich syndrome: report of an autosomal dominant variant. Blood 1996;87(11):4538-4543.##Knox-Macaulay H, Bashawri L, Davies KE. X linked recessive thrombocytopenia. J Med Gen 1993;30(11): 968-969.##He X, Zou R, Zhang B, You Y, Yang Y, Tian X. Whole Wiskott‑Aldrich syndrome protein gene deletion identified by high throughput sequencing. Mol Med Rep 2017;16(5):6526-6531.##Freson K. Clinical next generation sequencing to identify novel platelet disorders. Blood 2016;128(22):SCI-38.##Aldrich RA, Steinberg AG, Campbell DC. Pedigree demonstrating a sex-linked recessive condition chara-cterized by draining ears, eczematoid dermatitis and bloody diarrhea. Pediatrics 1954;13(2):133-139.##Derry JM, Ochs HD, Francke U. Isolation of a novel gene mutated in Wiskott-Aldrich syndrome. Cell 1994;78(4):635-644.##Jin Y, Mazza C, Christie JR, Giliani S, Fiorini M, Mella P, et al. Mutations of the Wiskott-Aldrich Syndrome Protein (WASP): hotspots, effect on transcription, and translation and phenotype/genotype correlation. Blood 2004;104(13):4010-4019.##Ramesh N, Geha R. Recent advances in the biology of WASP and WIP. Immunol Res 2009;44(1-3):99-111.##Sereni L, Castiello MC, Villa A. Platelets in Wiskott-Aldrich syndrome: victims or executioners? J Leukoc Biol 2018;103(3):577-590. ##Poulter NS, Pollitt AY, Davies A, Malinova D, Nash GB, Hannon MJ, et al. Platelet actin nodules are podosome-like structures dependent on Wiskott-Aldrich syndrome protein and ARP2/3 complex. Nat Commun 2015;6:7254.##Sabri S, Foudi A, Boukour S, Franc B, Charrier S, Jandrot-Perrus M, et al. Deficiency in the Wiskott-Aldrich protein induces premature proplatelet formation and platelet production in the bone marrow compartment. Blood 2006;108(1):134-140.##Massaad MJ, Ramesh N, Geha RS. Wiskott‐Aldrich syndrome: a comprehensive review. Ann N Y Acad Sci 2013;1285(1):26-43. ##Mantadakis E, Sawalle‐Belohradsky J, Tzanoudaki M, Kanariou M, Chatzimichael A, Albert MH. X‐linked thrombocytopenia in three males with normal sized platelets due to novel WAS gene mutations. Pediatr Blood Cancer 2014;61(12):2305-2306.##Sokolic R, Oden N, Candotti F. Assessment of immature platelet fraction in the diagnosis of Wiskott-Aldrich syndrome. Front Pediatr 2015;3:49.##Bastida JM, Del Rey M, Revilla N, Benito R, Perez-Andrés M, González B, et al. Wiskott-Aldrich syndrome in a child presenting with macrothrombocytopenia. Platelets 2017;28(4):417-420.##Derry JM, Ochs HD, Francke U. Isolation of a novel gene mutated in Wiskott–Aldrich syndrome. Cell 1994;78(4):635-644. ##Aldrich RA, Steinberg AG, Campbell DC. Pedigree demonstrating a sex-linked recessive condition chara-cterized by draining ears, eczematoid dermatitis and bloody diarrhea. Pediatrics 1954;13(2):133-139.##Remold-O’Donnell E, Rosen FS, Kenney DM. Defects in Wiskott-Aldrich syndrome blood cells. Blood 1996;87(7):2621-2631. ##Imai K, Nonoyama S, Ochs HD. WASP (Wiskott-Aldrich syndrome protein) gene mutations and phenotype. Curr Opin Allergy Clin Immunol 2003;3(6):427-436.##Yu H, Liu T, Meng W, Hou L. A novel WASP gene mutation in a Chinese boy with Wiskott-Aldrich syndrome. Int J Hematol 2010;92(2):271-275.##Wu H, Hu C, Dang D, Guo Y-J. A novel WASP gene mutation in a Chinese boy with Wiskott-Aldrich synd-rome. Indian J Hematol Blood Transfus 2014;30(1):353-355. ##Buchbinder D, Nadeau K, Nugent D. Monozygotic twin pair showing discordant phenotype for X-linked thrombocytopenia and Wiskott-Aldrich syndrome: a role for epigenetics? J Clin Immunol 2011;31(5):773-777.##

In Vitro Pre-validation of Gene Editing by CRISPR/Cas9 Ribonucleoprotein 2 2 Background: The CRISPR/Cas9 genome editing system is a powerful and simple gene editing method. The format of the CRISPR components is one of the important factors in targeting efficiency. Compared to plasmid or mRNA (IVTs) format, using the CRISPR/Cas9 system as Cas9–crRNA–tracrRNA RNP format is more efficient and rapid, especially in minimizing some of the pitfalls of CRISPR-mediated gene editing. In addition to efficient in vivo applications of the CRISPR RNP format in a variety of cell types and organisms, another advantage of this approach is usability for in vitro applications in which the crRNAs in the tracrRNA–crRNA structure guides the Mg2+-dependent RNAdirected DNA endonuclease to introduce double-strand breaks at specific sites in DNA. Methods: Here, Cas9–crRNA–tracrRNA RNP system was used to test the designed crRNAs for in vitro DNA cleavage by Cas9 protein in RAG1, RAG2 and IL2RG genes.  Results: The results of cleavage reveal theCas9–crRNA–tracrRNA RNP system is a rapid and efficient way to pre-validate the efficiency of CRISPR cleavage with crRNAs designed for RAG1, RAG2 and IL2RG genes. Conclusion: one step in vitro cleavage of DNA by CRISPR/Cas9 ribonucleoprotein complex can be used to pre-validate the functionality and relative efficiency of CRISPR system for targeting genes. 259 263 Maryam Mehravar Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Iran Abolfazl Shirazi Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECRResearch Institute of Animal Embryo Technology, Shahrekord University Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECRResearch Institute of Animal Embryo Technology, Shahrekord University IranIran Mohammad Mehdi Mehrazar Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR Iran Mahboobeh Nazari Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR Iran CRISPR/Cas9In vitro digestionRibonucleoprotein 10384.pdf Zhang F, Maeder ML, Unger-Wallace E, Hoshaw JP, Reyon D, Christian M, et al. High frequency targeted mutagenesis in Arabidopsis thaliana using zinc finger nucleases. Proc Natl Acad Sci USA 2010;107(26):12028-12033.##Zhang F, Cong L, Lodato S, Kosuri S, Church GM, Arlotta P. Efficient construction of sequence-specific TAL effectors for modulating mammalian transcription. Nat Biotechnol 2011;29(2):149-153.##Briggs AW, Rios X, Chari R, Yang L, Zhang F, Mali P, et al. Iterative capped assembly: rapid and scalable synthesis of repeat-module DNA such as TAL effectors from individual monomers. Nucleic Acids Res 2012;40 (15):e117-e117.##Carroll D, Morton JJ, Beumer KJ, Segal DJ. Design, construction and in vitro testing of zinc finger nucleases. Nat Protoc 2006;1(3):1329-1341.##Sanjana NE, Cong L, Zhou Y, Cunniff MM, Feng G, Zhang F. A transcription activator-like effector toolbox for genome engineering. Nature Protocols 2012;7(1):171.##Pennisi E. The CRISPR craze. Science 2013;341(6148):833-836.##Cong L, Ran FA, Cox D, Lin S, Barretto R, Habib N, et al. 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Cloning and Expression of B. mellitensis bp26 Gene in Lactococcus lactis as a Food Grade Vaccine 2 2 Background: Brucellosis is still an important health problem in under developing countries and researches for finding efficient vaccine are going on. Brucella melitensis (B. mellitensis) bp26 gene is a good candidate for brucellosis vaccine and investigations showed that Lactococcus lactis (L. lactis) with several positive characteristic are attractive for protein expression as a live delivery vectors. These fast growing bacteria need no aeration, are easy to handle, have no exotoxin, endotoxin and protease, so the cost of culturing is inexpensive. Methods: B. mellitensis bp26 gene was cloned in food grade pNZ 8149 vector and expressed in L. lactis NZ 3900. Results: Results showed that we can produce a food-grade recombinant L. lactis producing the B. melitensis BP26 protein.  Conclusion: In this study, for Future evaluation about ability of L. lactis as a live delivery vector, a food-grade recombinant L. lactis producing the B. melitensis BP26 protein was produced.  264 267 Maryam Azizpour Maghvan Department of Microbiology, Islamic Azad University, Arak Branch Department of Microbiology, Islamic Azad University, Arak Branch Iran Parvaneh Jafari Department of Microbiology, Islamic Azad University, Arak Branch Department of Microbiology, Islamic Azad University, Arak Branch Iran Seyyed Davood Hosseini Razi Vaccine and Serum Research Institute, Agricultural Research, Education & Extension Organization Razi Vaccine and Serum Research Institute, Agricultural Research, Education & Extension Organization Iran Ali mohammad Behrozikhah Razi Vaccine and Serum Research Institute, Agricultural Research, Education & Extension Organization Razi Vaccine and Serum Research Institute, Agricultural Research, Education & Extension Organization Iran BrucellosisExotoxinsLactococcus lactisVaccines 10385.pdf Avila-Calderón ED, Lopez-Merino A, Sriranganathan N, Boyle SM, Contreras-Rodríguez A. A history of the development of brucella vaccines. BioMed Res Int 2013;2013:743509.##Acharya KP, Kaphle K, Shrestha K, Bastuji BG, Smits HL. Review of brucellosis in Nepal. Int J Veterinary Sci Med 2016;4(2):54-62.##Basiri H, Akbari N, Azizpour M, Hossein SD, Behrozikhah AM, Eskandari S. Amplification, cloning and expression of Brucella melitensis bp26 gene (OMP28) isolated from Markazi province (Iran) and purification of Bp26 protein. Archives Razi Institute 2013;68(2):111-116.##Chaudhuri P, Prasad R, Kumar V, Gangaplara A. Recombinant OMP28 antigen-based indirect ELISA for serodiagnosis of bovine brucellosis. Mol Cell Probes 2010;24(3):142-145.##Gupta VK, Radhakrishnan G, Harms J, Splitter G. Invasive Escherichia coli vaccines expressing Brucella melitensis outer membrane proteins 31 or 16 or periplasmic protein BP26 confer protection in mice challenged with B. melitensis. Vaccine 2012;30(27):4017-4022.##Pontes DS, de Azevedo MS, Chatel JM, Langella P, Azevedo V, Miyoshi A. 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