Arak University Cell and Tissue Journal 2228-7035 8 1 2017 03 21 Investigation of Cell Cycle, Myogenic and Growth Factor Genes Expression in Ovine Primary Myoblast Cells by Follistatin Overexpression Investigation of Cell Cycle, Myogenic and Growth Factor Genes Expression in Ovine Primary Myoblast Cells by Follistatin Overexpression 1 11 26495 10.52547/JCT.8.1.1 FA M Nazari 1. Department of Animal Science, Faculty of Animal Science and Food Technology, Ramin Agriculture and Natural Resources University of Khuzestan, Ahvaz-Iran F Salabi Department of Venomous Animals and Anti-venom Production, Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Ahvaz-Iran L Du National Center for Molecular Genetics and Breeding of Animal, Chinese Academy of Agricultural Sciences, People’s Republic of China Journal Article 2017 07 22 Aim: The aim of this study was investigation of Follistatin gene over-expression effects on cell cycle, myogenic and growth factor genes expression of ovine primary myoblast (OPM) cells in lab that was done using AAV serotype 2 virus (rAAV2) carrying Follistatin (FST). Material and Methods: Obtained primary myoblasts cells from 60-day-old sheep fetuses were cultured in the growth and differentiation media. The optical density at 450 nm was measured as an indicator of proliferation in transfected cells with AAV serotype 2 (rAAV2) carrying Follistatin. Finally, Myogenic, Myo D, CDK2, Myf5, p57 and p21 genes expressions were measured using Real-Time qPCR. Results: The optical density (at 450 nm) was significantly increased in virus infected cells that shows Follistatin can increase proliferation. Follistatin proliferation significantly increased the expression of Akt1 and CDK2 genes and decreased the p21 expression gene under proliferation conditions. Real-Time results showed that the expression of myoincin, Myo D, Myf5, p57 and p21 genes increased under conditions of differentiation with Folistatin elevation. Conclusions: The results showed that proliferation of Follistatin increased the myoblast cells proliferation by p21 gene expressing and increasing the differentiation of myoblast cells by increasing the myogenic genes expression. Aim: The aim of this study was investigation of Follistatin gene over-expression effects on cell cycle, myogenic and growth factor genes expression of ovine primary myoblast (OPM) cells in lab that was done using AAV serotype 2 virus (rAAV2) carrying Follistatin (FST). Material and Methods: Obtained primary myoblasts cells from 60-day-old sheep fetuses were cultured in the growth and differentiation media. The optical density at 450 nm was measured as an indicator of proliferation in transfected cells with AAV serotype 2 (rAAV2) carrying Follistatin. Finally, Myogenic, Myo D, CDK2, Myf5, p57 and p21 genes expressions were measured using Real-Time qPCR. Results: The optical density (at 450 nm) was significantly increased in virus infected cells that shows Follistatin can increase proliferation. Follistatin proliferation significantly increased the expression of Akt1 and CDK2 genes and decreased the p21 expression gene under proliferation conditions. Real-Time results showed that the expression of myoincin, Myo D, Myf5, p57 and p21 genes increased under conditions of differentiation with Folistatin elevation. Conclusions: The results showed that proliferation of Follistatin increased the myoblast cells proliferation by p21 gene expressing and increasing the differentiation of myoblast cells by increasing the myogenic genes expression.

https://jct.araku.ac.ir/article_26495_7f9e4c8e1de05725d2b30298eda1f306.pdf

Arak University Cell and Tissue Journal 2228-7035 8 1 2017 03 21 Evaluation of myelin sheath and their related proteins (MAG, MOG, MBP and PLP) during cuprizone-induced demyelination in central nervous system of male C57BL/6 mice Evaluation of myelin sheath and their related proteins (MAG, MOG, MBP and PLP) during cuprizone-induced demyelination in central nervous system of male C57BL/6 mice 12 21 26500 10.52547/JCT.8.1.12 FA L N Sanadgo Department of Biology, Faculty of Science, Zabol University, Zabol, Iran 0000-0002-4509-5336 I M Komijan Department of Biology, Faculty of Science, Arak University, Arak, Iran, 38156-8-8349 Journal Article 2017 07 22 Aim: The aim of this study was to investigate of cuprizone time-dependent effects on MAG, MOG, MBP, and PLP proteins and demyelination process. Materials and Methods: The myelin degradation model was created with a five week diet containing 0.2% of cuprizone. The rats brain corpus callosum region was subjected to tissue and molecular evaluation at weeks 3, 4 and 5. At the end of the fifth week, behavioral studies were performed by open field test. Results: Results showed that the expression of myelin sheath proteins decreased time-dependently in comparison with the control group. Despite the overall reduction of these proteins in the third week, only PLP protein decreased significantly in this week comparing to control (p <0.01). Other proteins also showed a significant reduction in expression after the fourth week. Electron microscopy studies showed significant changes in the reduction of myelin diameter in comparison to the control group only after 4 weeks (p < 0.01). Behavioral studies also indicated that cuprizone was able to cause significant behavioral abnormalities in the animals from the beginning of the third week compared to the control group. Conclusion: This research was the first time to study the main myelin proteins at the same time, and the results show that the best time for accurate molecular, tissue and behavioral reviews are fourth and fifth weeks. Aim: The aim of this study was to investigate of cuprizone time-dependent effects on MAG, MOG, MBP, and PLP proteins and demyelination process. Materials and Methods: The myelin degradation model was created with a five week diet containing 0.2% of cuprizone. The rats brain corpus callosum region was subjected to tissue and molecular evaluation at weeks 3, 4 and 5. At the end of the fifth week, behavioral studies were performed by open field test. Results: Results showed that the expression of myelin sheath proteins decreased time-dependently in comparison with the control group. Despite the overall reduction of these proteins in the third week, only PLP protein decreased significantly in this week comparing to control (p <0.01). Other proteins also showed a significant reduction in expression after the fourth week. Electron microscopy studies showed significant changes in the reduction of myelin diameter in comparison to the control group only after 4 weeks (p < 0.01). Behavioral studies also indicated that cuprizone was able to cause significant behavioral abnormalities in the animals from the beginning of the third week compared to the control group. Conclusion: This research was the first time to study the main myelin proteins at the same time, and the results show that the best time for accurate molecular, tissue and behavioral reviews are fourth and fifth weeks.

https://jct.araku.ac.ir/article_26500_9f2da2656cb730d8886db16b9cf661b2.pdf

Arak University Cell and Tissue Journal 2228-7035 8 1 2017 03 21 Investigation of the Effect of Inhalation of Magnetic Iron Oxide Nanoparticles on the Expression of P53 in the Testis Tissue of Balb/C Mice by Immunohistochemically Staining Investigation of the Effect of Inhalation of Magnetic Iron Oxide Nanoparticles on the Expression of P53 in the Testis Tissue of Balb/C Mice by Immunohistochemically Staining 22 31 26497 10.52547/JCT.8.1.22 FA Z Nasiri Department of Biology, Faculty of Science, Esfahan University, Esfahan, Iran S.J Moshtaghian Department of Biology, Faculty of Science, Esfahan University, Esfahan, Iran F Esmaeili Department of Biology, Faculty of Science, Esfahan University, Esfahan, Iran Journal Article 2017 07 22 Aim: The present study was designed to investigate the effects of chitosan/poly vinyl alcohol scaffold incorporated by NRBE (neonatal rat brain extract) on neuronal differentiation of P19 EC (embryonal carcinoma) stem cells. Material and Methods: In order to induce neuronal phenotype, P19-derived embryonic carcinoma stem cells were cultured on a rat chitosan/poly (vinyl alcohol) scaffold with newborn rat brain extracts. To evaluate the survival potential, migration and differentiation of the three-dimensional culture cells, these cells were grafted to the developing central nervous system of the chick embryo. Finally, the transplanted cells were detected using specific staining and immunofluorescence methods. Results: Cresyl-Violet specific staining confirmed the neuronal phenotype of the transplantation cells. Also, the expression of specific neural protein, synaptophysin, was shown using the immunofluorescence process. Conclusion: The results of this study showed that P19 embryonic carcinoma stem cells can be used as a source of pluripotent cells in transplantation studies in order to investigate cellular and molecular aspects of developmental and cellular differentiation. Aim: The present study was designed to investigate the effects of chitosan/poly vinyl alcohol scaffold incorporated by NRBE (neonatal rat brain extract) on neuronal differentiation of P19 EC (embryonal carcinoma) stem cells. Material and Methods: In order to induce neuronal phenotype, P19-derived embryonic carcinoma stem cells were cultured on a rat chitosan/poly (vinyl alcohol) scaffold with newborn rat brain extracts. To evaluate the survival potential, migration and differentiation of the three-dimensional culture cells, these cells were grafted to the developing central nervous system of the chick embryo. Finally, the transplanted cells were detected using specific staining and immunofluorescence methods. Results: Cresyl-Violet specific staining confirmed the neuronal phenotype of the transplantation cells. Also, the expression of specific neural protein, synaptophysin, was shown using the immunofluorescence process. Conclusion: The results of this study showed that P19 embryonic carcinoma stem cells can be used as a source of pluripotent cells in transplantation studies in order to investigate cellular and molecular aspects of developmental and cellular differentiation.

https://jct.araku.ac.ir/article_26497_a66dbabd793deed367030aadfcfc59b9.pdf

Arak University Cell and Tissue Journal 2228-7035 8 1 2017 03 21 Study the effects of some acute environmental conditions on recombinant Hsp70 protein expression in Rutilus Frissi Kutum liver in E. coli Study the effects of some acute environmental conditions on recombinant Hsp70 protein expression in Rutilus Frissi Kutum liver in E. coli 32 41 26496 10.52547/JCT.8.1.32 FA O Saberi Department of Biology, University of Guilan, University Campus 2, Rasht, Iran H Ghafoor Department of Biology, Faculty of science, University of Guilan, Rasht, Iran Journal Article 2017 03 21 Aim: The aim of the present study was to investigate the optimal expression of Hsp70 recombinant Rutilus FrissiKutum liver in E. coli. Material and methods: To evaluating of expression level of recombinant Hsp70, transformed E. coli were cultured in LB medium at 2, 4, 8, 12, 24, 36 and 48 hours, 20, 25, 30, 37 and 50 °C, extreme pH (5 and 9) and in present of heavy metals e.g. mercury, cobalt, iron, zinc and etc. Expression of recombinant Hsp70 protein was determined by SDS-PAGE 12% analysis of E. coli extracts followed by staining with Coomassie Blue. Results: Optimum expression of recombinant Hsp70 was obtained at 4 and 8 h, 37 °C and pH 5. Additionally, the maximum and minimum expression of this protein was achieved in present of manganese and cobalt respectively. The results further indicate that the survival of E. coli with Hsp70 was enhanced compared to the control cells. Conclusion: All results reveal that transformed bacteria are able to survive against extreme environmental conditions compared to control sample, because of having recombinant Hsp70. This heat-shock protein protects vital proteins from denaturing by its chaperone property. So, growth condition described in this study can be used for optimizing production of recombinant Hsp70 in E. coli host. Aim: The aim of the present study was to investigate the optimal expression of Hsp70 recombinant Rutilus FrissiKutum liver in E. coli. Material and methods: To evaluating of expression level of recombinant Hsp70, transformed E. coli were cultured in LB medium at 2, 4, 8, 12, 24, 36 and 48 hours, 20, 25, 30, 37 and 50 °C, extreme pH (5 and 9) and in present of heavy metals e.g. mercury, cobalt, iron, zinc and etc. Expression of recombinant Hsp70 protein was determined by SDS-PAGE 12% analysis of E. coli extracts followed by staining with Coomassie Blue. Results: Optimum expression of recombinant Hsp70 was obtained at 4 and 8 h, 37 °C and pH 5. Additionally, the maximum and minimum expression of this protein was achieved in present of manganese and cobalt respectively. The results further indicate that the survival of E. coli with Hsp70 was enhanced compared to the control cells. Conclusion: All results reveal that transformed bacteria are able to survive against extreme environmental conditions compared to control sample, because of having recombinant Hsp70. This heat-shock protein protects vital proteins from denaturing by its chaperone property. So, growth condition described in this study can be used for optimizing production of recombinant Hsp70 in E. coli host.

https://jct.araku.ac.ir/article_26496_1697fe5eb0141dca1379090a4d766cc7.pdf

Arak University Cell and Tissue Journal 2228-7035 8 1 2017 03 21 Effect of Glycyrriza glabra L. extract on ODC1 gene expression and cell proliferation in human breast cancer cells Effect of Glycyrriza glabra L. extract on ODC1 gene expression and cell proliferation in human breast cancer cells 42 51 26502 10.52547/JCT.8.1.42 FA A Safipour Afshar Department of Biology, Neyshabur Branch, Islamic Azad University, Neyshabur, Iran F Saeid Nematpour Department of Biology, Neyshabur Branch, Islamic Azad University, Neyshabur, Iran M Lakzian Department of Biology, Neyshabur Branch, Islamic Azad University, Neyshabur, Iran Journal Article 2017 07 22 Aim: Product of ODC1 gene is key enzyme in polyamines synthesis that in diverse cancers their content increases. In the present study the effects of root extract of licorice on the ODC1gene expression and cell viability of two human breast cancer cell lines (MCF-7 and MDA-MB-231) and non-cancer cell line (MCF-10A) were evaluated. Material and Methods: The cell lines were subjected to increasing doses of the extract ranging from 0-200 µg ⁄ml. Cytotoxic effect was assessed using the MTT assay. Expression of ornithine decarboxylase 1 gene was analyzed by Real Time PCR. Results: In cancerous cell lines mortality increases significantly by a concentration-dependent manner. Moreover, a marked decrease observed in the expression of ODC1 gene in cancer cell lines. In comparison of two cancer cell lines, MDA-MB-231cell line more affected by licorice root extract. Conclusion: The result of our study highlights the potential inﬂuences of Glycyrrhiza glabra extract on ODC1 gene expression in breast cancer cells and its relation to inhibition of cancer cell growth. Aim: Product of ODC1 gene is key enzyme in polyamines synthesis that in diverse cancers their content increases. In the present study the effects of root extract of licorice on the ODC1gene expression and cell viability of two human breast cancer cell lines (MCF-7 and MDA-MB-231) and non-cancer cell line (MCF-10A) were evaluated. Material and Methods: The cell lines were subjected to increasing doses of the extract ranging from 0-200 µg ⁄ml. Cytotoxic effect was assessed using the MTT assay. Expression of ornithine decarboxylase 1 gene was analyzed by Real Time PCR. Results: In cancerous cell lines mortality increases significantly by a concentration-dependent manner. Moreover, a marked decrease observed in the expression of ODC1 gene in cancer cell lines. In comparison of two cancer cell lines, MDA-MB-231cell line more affected by licorice root extract. Conclusion: The result of our study highlights the potential inﬂuences of Glycyrrhiza glabra extract on ODC1 gene expression in breast cancer cells and its relation to inhibition of cancer cell growth.

https://jct.araku.ac.ir/article_26502_c1020880214e7bbf222d0069b7c77cb7.pdf

Arak University Cell and Tissue Journal 2228-7035 8 1 2017 03 21 Influence of Occupational Exposures on Sperm Parameters and Chromatin Structure Influence of Occupational Exposures on Sperm Parameters and Chromatin Structure 52 68 26503 10.52547/JCT.8.1.52 FA L Babakhah Department of Reproductive Biotechnology, Reproductive Biomedicine Research Center, Royan Institute for Biotechnology, ACECR, Isfahan, Iran I L Azad Department of Reproductive Biotechnology, Reproductive Biomedicine Research Center, Royan Institute for Biotechnology, ACECR, Isfahan, Iran M Arbabian Department of Reproductive Biotechnology, Reproductive Biomedicine Research Center, Royan Institute for Biotechnology, ACECR, Isfahan, Iran M Tavalaee Department of Reproductive Biotechnology, Reproductive Biomedicine Research Center, Royan Institute for Biotechnology, ACECR, Isfahan, Iran M Bahadorani Department of Biology, Falavarjan Branch, Islamic Azad University, Isfahan, Iran MH Nasr- Esfahani Isfahan Fertility and Infertility Center, Isfahan, Iran Journal Article 2017 07 22 Aim: This study examines the sperm parameters (concentration, mobility and morphology) and the state of sperm DNA in infertile men exposed to various occupational exposures. Material and Methods: This study was performed on 152 men who referred to Isfahan Fertility and Infertility Center )spring 2014 to autumn 2016( for diagnostic purposes. After an interview with all the participants, their semen samples were collected for assessment of sperm parameters and sperm chromatin health, based on the WHO protocol and the TUNEL method, respectively. Results: The results showed that in men with prolonged sitting, high physiological activity and with exposure to chemical compounds, they had lower sperm motility and higher sperm DNA damage than the presented criteria by previous studies. In addition, in men with DNA damage higher than 10%, there was a significant decrease in sperm parameters. Conclusion: Occupational exposure has negative effects on sperm parameters and sperm chromatin health, but more studies are needed to prove these results. Aim: This study examines the sperm parameters (concentration, mobility and morphology) and the state of sperm DNA in infertile men exposed to various occupational exposures. Material and Methods: This study was performed on 152 men who referred to Isfahan Fertility and Infertility Center )spring 2014 to autumn 2016( for diagnostic purposes. After an interview with all the participants, their semen samples were collected for assessment of sperm parameters and sperm chromatin health, based on the WHO protocol and the TUNEL method, respectively. Results: The results showed that in men with prolonged sitting, high physiological activity and with exposure to chemical compounds, they had lower sperm motility and higher sperm DNA damage than the presented criteria by previous studies. In addition, in men with DNA damage higher than 10%, there was a significant decrease in sperm parameters. Conclusion: Occupational exposure has negative effects on sperm parameters and sperm chromatin health, but more studies are needed to prove these results.

https://jct.araku.ac.ir/article_26503_a6d01d2e4f6cc4bb41090e25bcc1fdcf.pdf

Arak University Cell and Tissue Journal 2228-7035 8 1 2017 03 21 Antibacterial effect of Modified Zeolite with Schiff base Antibacterial effect of Modified Zeolite with Schiff base 69 76 26504 10.52547/JCT.8.1.69 FA M Zendehdel Chemistry Department, Faculty of Sciences, Arak University, Arak 38156-8-8349, Iran Z Mortezaei Chemistry Department, Faculty of Sciences, Arak University, Arak 38156-8-8349, Iran M Yusefi Nano biotechnology Research Center, Avicenna Research Institute, Tehran, Iran Journal Article 2017 07 22 - Aim: In this study, solid hybrid material from the Schiff base complex was prepared inside and on the surface of NaY zeolite, and then their antibacterial properties against different bacteria (gram-positive and gram negative bacteria) was investigated. Material and Methods: Antibacterial compounds were first prepared by sequential synthesis. So that Schiff base complexes of intermediate metals are used in two ways, one way through the vessel's method inside the bottle, inside the NaY zeolite cavities, and again by the factorizing, which is the same as the modification of the pitted compound surface, on NaOH zeolite were loaded. In the second part, the antibacterial activity of the synthesized compounds in the laboratory was investigated against gram-positive (Bacillus subtilis and Staphylococcus aureus) and gram negative bacteria (Pseudomonas aeruginosa and Escherichia coli). Results: The results show that solid hybrid materials from Schiff base complexes and Zeolite NaY has fine antibacterial activity (against gram negative bacteria such as Pseudomonas aeruginosa and Escherichia coli) compared with Gentamicin and Nalidixic acid as standard drugs. So, fine inhibition on bacterial growth till 24 hours. Conclusion: The solid hybrid materials from Schiff base complexes and NaY Zeolite show fine inhibition on bacterial growth in comparison with standard drugs. In particular, these compounds can be used several times without losing their properties and making them industrially economical. - Aim: In this study, solid hybrid material from the Schiff base complex was prepared inside and on the surface of NaY zeolite, and then their antibacterial properties against different bacteria (gram-positive and gram negative bacteria) was investigated. Material and Methods: Antibacterial compounds were first prepared by sequential synthesis. So that Schiff base complexes of intermediate metals are used in two ways, one way through the vessel's method inside the bottle, inside the NaY zeolite cavities, and again by the factorizing, which is the same as the modification of the pitted compound surface, on NaOH zeolite were loaded. In the second part, the antibacterial activity of the synthesized compounds in the laboratory was investigated against gram-positive (Bacillus subtilis and Staphylococcus aureus) and gram negative bacteria (Pseudomonas aeruginosa and Escherichia coli). Results: The results show that solid hybrid materials from Schiff base complexes and Zeolite NaY has fine antibacterial activity (against gram negative bacteria such as Pseudomonas aeruginosa and Escherichia coli) compared with Gentamicin and Nalidixic acid as standard drugs. So, fine inhibition on bacterial growth till 24 hours. Conclusion: The solid hybrid materials from Schiff base complexes and NaY Zeolite show fine inhibition on bacterial growth in comparison with standard drugs. In particular, these compounds can be used several times without losing their properties and making them industrially economical.

https://jct.araku.ac.ir/article_26504_404d52bf50ab4f86fbf54376d0bcf0dd.pdf

Arak University Cell and Tissue Journal 2228-7035 8 1 2017 03 21 Purification of Influenza A (H1N1) Recombinant Nucleoprotein , Preparation and Purification of its Polyclonal Antibody in Rabbits Purification of Influenza A (H1N1) Recombinant Nucleoprotein , Preparation and Purification of its Polyclonal Antibody in Rabbits 77 87 26498 10.52547/JCT.8.1.77 FA I M Sharaf Department of Poultry Science, College of Agriculture, Tarbiat Modares University, Tehran, Iran. Z Asgariazadeh - Department of Poultry Science, College of Agriculture, Tarbiat Modares University, Tehran, Iran. MA Karimi Torshizi Department of Poultry Science, College of Agriculture, Tarbiat Modares University, Tehran, Iran. Journal Article 2017 07 22 - Aim: This study was carried out to investigate the application of different magnetic intensities on the sperm freezing medium. Material and methods: In this experiment, the magnetic field induction of 0, 2000, 4000 and 6000 Gauss was used to determine the optimum intensity during the freeze-thawing process. Semen samples were collected from 10 pieces of 24-week-old Ross 308 broiler chickens using abdominal massage technique. In this study, the experimental treatments were: 1) ordinary diluent, 2) G2000-intensive magnetic diluent, 3) G4000-intensive magnetic diluent, and G6000-intensive magnetic diluent. The used diluent in this study was the Lake diluent. This diluent was mixed with 1% soy lecithin and 5% glycerol, and then on the basis of the experimental treatments, they encountered different magnetic intensities. Diluted sperm was frozen based on each treatment, after cooling in a 0.25 ml straws. After freezing some sperm parameters such as total and progressive motilities, morphology, membrane integrity, MDA production, apoptosis status and mitochondrial activity were evaluated. Results: Results showed that different magnetic fields had no significant effect on total and progressive mobilities, membrane integrity, morphology, lipid peroxidation, acrosome integrity and apoptosis mechanisms of rooster sperm. Just mitochondrial membrane potential in the control group was higher than other groups (60.5%, p < 0.05). Conclusion: It seems that exposure of rooster semen cryopreservation media with magnetic field cannot be efficient for quality of sperm after thawing. - Aim: This study was carried out to investigate the application of different magnetic intensities on the sperm freezing medium. Material and methods: In this experiment, the magnetic field induction of 0, 2000, 4000 and 6000 Gauss was used to determine the optimum intensity during the freeze-thawing process. Semen samples were collected from 10 pieces of 24-week-old Ross 308 broiler chickens using abdominal massage technique. In this study, the experimental treatments were: 1) ordinary diluent, 2) G2000-intensive magnetic diluent, 3) G4000-intensive magnetic diluent, and G6000-intensive magnetic diluent. The used diluent in this study was the Lake diluent. This diluent was mixed with 1% soy lecithin and 5% glycerol, and then on the basis of the experimental treatments, they encountered different magnetic intensities. Diluted sperm was frozen based on each treatment, after cooling in a 0.25 ml straws. After freezing some sperm parameters such as total and progressive motilities, morphology, membrane integrity, MDA production, apoptosis status and mitochondrial activity were evaluated. Results: Results showed that different magnetic fields had no significant effect on total and progressive mobilities, membrane integrity, morphology, lipid peroxidation, acrosome integrity and apoptosis mechanisms of rooster sperm. Just mitochondrial membrane potential in the control group was higher than other groups (60.5%, p < 0.05). Conclusion: It seems that exposure of rooster semen cryopreservation media with magnetic field cannot be efficient for quality of sperm after thawing.

https://jct.araku.ac.ir/article_26498_729f2c7eeeab66f50f4ab3677539a585.pdf

Arak University Cell and Tissue Journal 2228-7035 8 1 2017 03 21 Investigating of protective effects of Nurr1 and GDNF on dopaminergic neural cell line SH-SY5Y against neuro-inflammation and toxicity of 6-OHDA Investigating of protective effects of Nurr1 and GDNF on dopaminergic neural cell line SH-SY5Y against neuro-inflammation and toxicity of 6-OHDA 87 108 26501 10.52547/JCT.8.1.87 FA M Rasoolnezhad Master graduate in cellular and molecular biology, National Institute of Genetic Engineering and Biotechnology, Tehran. M Gardaneh Dept of Stem cells and Regenerative Medicine Group, Faculty of Molecular Genetics, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran Sabouni F Dept of Molecular Medicine, Faculty of Biochemistry, National Institute of Genetic Engineering and Biotechnology, Tehran, , Iran Journal Article 2017 07 22 - Aim: In this study Nurr1 and GDNF, due to their anti-inflammatory and regenerative effects and Nurr1-mediated regulation of GDNF receptor (Ret) expression, were selected to protect dopaminergic SH-SY5Y cell line against neuroinflammation and toxicity caused by 6-OHDA. Material and Methods: Recombinant lentiviral vectors carrying Nurr1 and GDNF genes were prepared and transdused to SH-SY5Y and astrocytoma (1321N1) cell lines respectively. Also HEK-293T cells were transfected with plasmid carrying GDNF to overexpress this factor; condition media of transduced astrocytoma and transfected HEK-293T cells were collected and stored. Next, overexpression of mentioned factors was demonstrated by RT-PCR. On the other hand, microglial cells were isolated from neonatal rat brains and induced with LPS to produce neuroinflammatory factors; Inducible expression of them was demonstrated by Griess test and RT-PCR. Condition media of microglia was collected and saved. Finally, SH-SY5Y cells overexpressing Nurr1 were treated with condition media of transduced astrocytoma / transfected HEK-293T and then with condition media of LPS-induced microglia or 6-OHDA toxin. Results: data from MTT assay showed, SH-SY5Y cells overexpressing Nurr1 or pretreated with GDNF are more resistant to toxicity caused by neuroinflammation and 6-OHDA. Also Nurr1 and GDNF have cooperative effects and give more protection to dopaminergic cells. Conclusion: Nurr1 and GDNF each have protective effects on dopaminergic neural cells against inflammatory factors or 6-OHDA. Also they synergize with each other leading to more protection for dopaminergic neural cells. - Aim: In this study Nurr1 and GDNF, due to their anti-inflammatory and regenerative effects and Nurr1-mediated regulation of GDNF receptor (Ret) expression, were selected to protect dopaminergic SH-SY5Y cell line against neuroinflammation and toxicity caused by 6-OHDA. Material and Methods: Recombinant lentiviral vectors carrying Nurr1 and GDNF genes were prepared and transdused to SH-SY5Y and astrocytoma (1321N1) cell lines respectively. Also HEK-293T cells were transfected with plasmid carrying GDNF to overexpress this factor; condition media of transduced astrocytoma and transfected HEK-293T cells were collected and stored. Next, overexpression of mentioned factors was demonstrated by RT-PCR. On the other hand, microglial cells were isolated from neonatal rat brains and induced with LPS to produce neuroinflammatory factors; Inducible expression of them was demonstrated by Griess test and RT-PCR. Condition media of microglia was collected and saved. Finally, SH-SY5Y cells overexpressing Nurr1 were treated with condition media of transduced astrocytoma / transfected HEK-293T and then with condition media of LPS-induced microglia or 6-OHDA toxin. Results: data from MTT assay showed, SH-SY5Y cells overexpressing Nurr1 or pretreated with GDNF are more resistant to toxicity caused by neuroinflammation and 6-OHDA. Also Nurr1 and GDNF have cooperative effects and give more protection to dopaminergic cells. Conclusion: Nurr1 and GDNF each have protective effects on dopaminergic neural cells against inflammatory factors or 6-OHDA. Also they synergize with each other leading to more protection for dopaminergic neural cells.

https://jct.araku.ac.ir/article_26501_a4f87e21ec472bbff8435588d5b08a8f.pdf