2024-03-29T09:52:09+04:30 http://ibj.pasteur.ac.ir/browse.php?mag_id=54&slc_lang=en&sid=1
54-512 2024-03-29 10.1002
Iranian Biomedical Journal IBJ 1028-852X 2008-823X 10.61186/ibj 2004 8 2 A Novel Vector for Expression/Secretion of Properly Folded Eukaryotic Proteins: a Comparative Study on Cytoplasmic and Periplasmic Expression of Human Epidermal Growth Factor in E. coli Mina Ebrahimi-Rad minebrad@yahoo.com Morteza Azarnoosh Aftab Bashir Vladimir V. Bakayev Expression of eukaryotic proteins in E. coli often results in their aggregation. Proper folding and solubility of therapeutical proteins are the pre-requisite for their bioactivity. This is not achieved in cytoplasmic expression in E. coli because of the absence of disulfide bonds formation. A novel expression/secretion vector was constructed which exploited β-lactamase signal sequence to translocate processed and soluble proteins into the periplasm of cells. Secretion of model proteins, β-lactamase and human Epidermal Growth Factor (hEGF) in M15/pSB and M15/pSE systems respectively, was confirmed by SDS-PAGE analysis and bioactivity assay. Secreted hEGF was found to be identical to authentic protein, in size, N-terminal amino acid sequence, biological activity and Western-blotting. The radioimmunoassay revealed 10-fold-higher level of hEGF expression in M15/pSE secretion system compared to that of cytoplasmic expression of the protein. The properly processed and in vivo folded hEGF demonstrated high solubility and bioactivity. The data obtained, evidenced in favor of M15/pSE expression system, which might be suitable to produce the small eukaryotic disulfide-bonded proteins for therapeutical applications or structural studies Protein expression Secretion β-lactamase Human Epidermal Growth Factor (hEGF) Signal sequence 2004 4 01 51 61 http://ibj.pasteur.ac.ir/article-1-512-en.pdf
54-513 2024-03-29 10.1002
Iranian Biomedical Journal IBJ 1028-852X 2008-823X 10.61186/ibj 2004 8 2 Human Sperm Aster Formation and Chromatin Configuration in Rabbit Oocytes Following Intracytoplasmic Sperm Injection Using a Piezo-Micromanipulator Fardin Amidi Farid Abolhassani Yukihiro Terada Azim Hedayatpour Marefat Ghaffari Hoshang Rafigdust Sou-ichi Nakamura Kunihiro Okamura In human fertilization, the sperm centrosome nucleates a radial array of microtubules called the sperm aster. The sperm aster is responsible for apposition of male and female pronuclei, and later gives rise to the first meiotic spindle. The objective of this study was to determine microtubule assembly and chromatin configuration in rabbit oocytes following intracytoplasmic injection with human sperm by piezo-driven pipette. Oocytes were collected from superovulated dose 14-15 h after hCG injection and were fertilized by injection of a single human sperm into the ooplasm of each oocyte without additional activation treatment. Four hours post heterologous intracytoplamic sperm injection (ICSI), rabbit eggs were fixed and microtubule organization and chromatin configuration were examined by immunofluorescence microscopy. In unfertilized oocytes, microtubules were present only in the metaphase-arrested second meiotic spindle. Following human sperm injection, an aster of microtubules formed adjacent to the sperm head, around mid-piece, and sperm aster was enlarged and assembled around male and female pronuclei. During pronuclear centration, male and female pronuclei were surrounded by a microtubule array without nucleation sites. With fertile human sperm, the sperm aster formation rate was 54.6%. From our data we concluded that human spermatozoa can be injected successfully into rabbit oocytes, resulting in a reasonable survival rate, and that rabbit oocytes provide a reliable tools for assessing human sperm centrosomal function using the Piezo-ICSI system Centrosome Fertilization Intracytoplamic sperm injection (ICSI) Rabbit eggs 2004 4 01 63 68 http://ibj.pasteur.ac.ir/article-1-513-en.pdf
54-514 2024-03-29 10.1002
Iranian Biomedical Journal IBJ 1028-852X 2008-823X 10.61186/ibj 2004 8 2 Effect of Acute and Chronic Heat Exposure on Frequency of EEG Components in Different Sleep-Wake State in Young Rats Rakesh Kumar Sinha rksinha_res@rediffmail.com Amit Kumar Ray The recent literatures indicate that central nervous system (CNS) is highly vulnerable to systemic hyperthermia induced by whole body heating on conscious animals. In the present study, cerebral electrical activity or EEG (electroencephalogram) following exposure to high environmental heat has been studied in moving rats. Rats were divided into three group (i) acute heat stress-subjected to a single exposure for four hours in the BOD (Biological Oxygen Demand) incubator at 38°C (ii) chronic heat stress-exposed for 21 days daily for one hour in the incubator at 38°C, and (iii) handling control groups. EEG signals were recorded on paper along with in digital form on computer hard disk just after the heat exposure from acute stressed rats and on 22nd day from chronic stressed rats. EOG (electrooculogram) and EMG (electromyogram) were also recorded along with EEG to differentiate the different sleep-wake states. Following up, for each group of rats, power spectrum of the EEG for each sleep-wake states, and for all four hours of recording in two-second epochs were calculated and analyzed. The higher frequency components of the EEG power spectrum (b2) was found very sensitive to hot environment and changed significantly in all three sleep-wake states in comparison to the control rats following acute as well as chronic exposure to heat stress. This report demonstrates that the cortical EEG are sensitive to environmental heat and alterations in EEG frequencies in different state of mental consciousness due to high heat can be differentiated efficiently by EEG power spectrum analysis Acute heat stress Chronic heat stress Electroencephalogram Sleep-wake state Power spectrum 2004 4 01 69 75 http://ibj.pasteur.ac.ir/article-1-514-en.pdf
54-515 2024-03-29 10.1002
Iranian Biomedical Journal IBJ 1028-852X 2008-823X 10.61186/ibj 2004 8 2 Assessment of Epidermal Growth Factor (EGF) Effects on Development of Vitrified Mouse Morulae to the Blastocyst Stage Seyed Morteza Koruji Mansoureh Movahedin Mojtaba Rezazadeh Valojerdi Many investigators are interested in finding the new cultural systems that can support the in vitro development of pre-implantation embryos better. Previous studies suggested that growth factors such as epidermal growth factor (EGF) are important in pre-implantation embryo development and implantation process. On the other hand, it is very important to support post thaw development of frozen embryos. The purpose of this study was to determine if the developmental potential of mouse morulae survived after vitrification could be increased using medium containing EGF. Mouse morulae were divided into vitrified and non-vitrified groups. Vitrification procedure was carried out using a combination of 40% ethylene glycol, 30% ficoll and 0.5 M sucrose (EFS40) as cryoprotectant. The embryos were warmed rapidly using 0.5 M sucrose. The survived embryos were cultured either on T6 or T6+EGF media. Accordingly, the embryos of the non-vitrified group were also cultured. The developmental rates in all groups were daily recorded and compared statistically using Chi-square test. The results showed that after 4 days of culture, the developmental potential of non-vitrified embryos cultured on T6+EGF was significantly increased. There was no significant difference between vitrified embryos cultured on T6 and T6+EGF media. In conclusion, the developmental potential of vitrified-warmed embryos does not increase in the medium containing EGF, even though there was significant increased developmental potential of non-vitrified embryos after culture on medium containing EGF. It is needed to do more study about the changes which will probably happen on the embryo EGF receptors following vitrification Vitrification Growth factor Mouse embryo 2004 4 01 77 82 http://ibj.pasteur.ac.ir/article-1-515-en.pdf
54-516 2024-03-29 10.1002
Iranian Biomedical Journal IBJ 1028-852X 2008-823X 10.61186/ibj 2004 8 2 Extraction of the Outer Membrane Proteins of H. pylori and Evaluation of Their Presence in Stool of the Infected Individuals Ali Sheikhian Zuhair M. Hassan hassan_zm@yahoo.co.uk Ali Mustafaie Fazel Shokri Reza Malekzadeh Farideh Siavoshi Helicobacter pylori infections are accompanied with the release of some antigenic material into the stool. Outer membranes proteins (OMP) are among the major antigens of bacteria, which in comparison with other bacterial constituents have less cross-reactivity. In this study, OMP of H. pylori were isolated from 11 clinical isolates and rabbit antiserum against them was used to detect possible antigens of H. pylori, which are released into the stool. We used immunoblotting and affinity chromatography to detect such antigens in fecal antigenic extracts of infected individuals. By immunoblotting, we were able to detect a 26 kDa band under reducing and non-reducing conditions, but many more antigens (at least 5 antigens with molecular weights of about 14, 26, 52, 57.5 and 66 kDa) were isolated by affinity chromatography. The 26 kDa antigen had a higher concentration and is seen in nearly all positive samples. Since the 26 kDa antigen is detectable by these two techniques, we suggest that this antigen is one of the major antigens of H. pylori which is released into the stool and can be considered as a candidate diagnostic antigen to be used in diagnostic kit development H. pylori Outer membrane proteins (OMP) Stool SDS-PAGE Affinity chromatography 2004 4 01 83 88 http://ibj.pasteur.ac.ir/article-1-516-en.pdf
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Iranian Biomedical Journal IBJ 1028-852X 2008-823X 10.61186/ibj 2004 8 2 Molecular Evidence on Changing Pattern of Mixed Plasmodium falciparum and P. vivax Infections during Year-Round Transmission of Malaria in Chahbahar, Iran Sedigheh Zakeri zakeris@yahoo.com Parin Naimi Mohammad Zare Mehdi Zand Haghighi Navid Dinparast-Djadid Mixed malaria infections, Plasmodium falciparum and P. vivax, are suspected to occur at a greater frequency than is detected by conventional light microscopy. In order to determine the year round pattern of transmission and the frequency of mixed infections in malaria endemic area, we carried out a prospective comparison of diagnosis by conventional light microscopy and nested PCR in Chahbahar district, south-eastern part of Iran. Out of 280 Giemsa-stained slides, 158 (56.42%) were identified as having only P. vivax and 89 (31.78%) were P. falciparum infection by microscopy. Only eight slides (2.8%) were interpreted as having mixed P. vivax-P. falciparum infections and 25 (8.9%) were negative. Comparing to the microscopy results, the PCR detected 33 more mixed infections. These results showed that the number, of mixed infections was increased during April to September and reduced after September, although malaria cases with only P. falciparum were increased. The possibility that malaria patients in Chahbahar district may have undetected mixed infections during first peak of transmission should be kept in mind because of the specific therapy required both for P. falciparum and for radical cure of P. vivax Malaria P. falciparum P. vivax Mixed infections PCR 2004 4 01 89 93 http://ibj.pasteur.ac.ir/article-1-517-en.pdf
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Iranian Biomedical Journal IBJ 1028-852X 2008-823X 10.61186/ibj 2004 8 2 Partial Purification of a Potent Immunosuppressive Factor Excreted from Leishmania major Promastigote and Amastigote Mohsen Abolhassani mabolhassani@yahoo.com Haideh Darabi Recent scientific evidence indicates that distinct patterns of susceptibility in BALB/c mice to Leishmania major infection are attributable to the differential expansion of distinct CD4+ T-cell subsets and their cytokines production. Production of the Th1 cytokine IFN-g is associated with resistance, whereas production of the Th2 cytokines IL-4 and IL-10 are associated with extreme susceptibility. The major host immune defense mechanism against Leishmania is activation of macrophages by INF-γ derived from T cells. The inability of susceptible hosts to mount the immune response necessary to activate macrophage and destroy the parasites may be due to the parasite-specific proteins that are able to suppress the immune system. In the present study, we have semi-purified the excreted antigens of Leishmania major promastigote and amastigote by column chromatography. The isolated fraction showed a potent immunosuppressive activity on normal BALB/c mice lymphocytes stimulated with mitogens. Fifteen microgram of the isolated fraction caused 81% suppression of lymphocyte proliferation. These data may suggest that the parasite by secreting immunosuppressive factor down regulate the immune system and as a result survive in the body Leishmania major Amastigote Promastigote Immunosuppressive Factor Purification 2004 4 01 95 99 http://ibj.pasteur.ac.ir/article-1-518-en.pdf
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Iranian Biomedical Journal IBJ 1028-852X 2008-823X 10.61186/ibj 2004 8 2 Collagen as Adherent Substratum and Inducer of Dorsal Root Ganglia Outgrowth Massoumeh Firouzi Farzaneh Sabouni Abed-Ali Ziaee Mohammad Taghikhani Neurite outgrowth from dorsal root ganglion (DRG) explants is a method of evaluating neurotrophic activity of growth factors. When complete medium containing collagen was supplemented with nerve growth factor (NGF) DRG outgrowth was observed after 18 h. In the absence of NGF and in the presence of collagen, the DRG outgrowth took place after 72 h. In wells not supplemented with collagen gel in substratum, no DRG outgrowth was observed. Partially, DRG differentiation was observed in the presence of NGF. In the absence of NGF and collagen, there was no DRG outgrowth detected. It seems that, in some circumstances, cells degenerated by DRG may be an indication of an apoptosis phenomenon. Therefore, we suggested that collagen as a substratum is more effective than NGF Collagen Adherent substratum Dorsal root ganglia Culture Neurite 2004 4 01 101 105 http://ibj.pasteur.ac.ir/article-1-519-en.pdf
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Iranian Biomedical Journal IBJ 1028-852X 2008-823X 10.61186/ibj 2004 8 2 Cellobiose Dehydrogenase Production by the Genus Cladosporium Masoomeh Shams Ghahfarokhi shamsm@modares.ac.ir Ali Fazli Abbas Lotfi Mehdi Razzaghi Abyaneh Cellobiose dehydrogenase (CDH EC.1.1.5.1) is an extracellular enzyme that mainly produced by wood-degrading fungi. It oxidizes cellobiose to cellobionolactone using a wide spectrum of electron acceptors. The key roles of CDH in growth, metabolism, and some other important cellular processes such as cellulose degradation in fungi have been noted. Since the demands for finding new sources of CDH among different organisms have been dramatically increased, this study was focused on the presence of CDH in the genus Cladosporium as a well-known cellulolytic fungus. Twenty strains of Cladosporium isolated from soil samples from different geographical origin were evaluated for CDH-producing ability. The early screening of the fungus by zymogram method revealed the presence of CDH as an extracellular form in all of the examined isolates. Submerged cultivation of the best producer of CDH (selected from initial screening) on a specific medium showed the maximum amounts of enzyme produced in shaking cultures with pH 4.5 at 28ºC for a 14-day period. The enzyme activity was determined in the range of 27.83 to 1284.84 unit/mg protein among the isolates. Our observations show that Cladosporium isolates with high CDH producing ability i.e. isolates No. 10 and No. 18 can be used as selective candidates for large-scale production of this industrially important enzyme in further research programs. This is the first documented report on the presence of CDH in the fungus Cladosporium Cladosporium Cellobiose dehydrogenase Production Screening 2004 4 01 107 111 http://ibj.pasteur.ac.ir/article-1-520-en.pdf