Author/Authors :
Piri, Hossein qazvin university of medical sciences - Cellular and Molecular Research Center, School of Medicine - Department of Biochemistry and Genetics, ايران , Kazemi, Bahram shahid beheshti university of medical sciences - Cellular and Molecular Biology Research Center, School of Medicine - Department of Biotechnology, تهران, ايران , Khodadadi, Iraj hamadan university of medical sciences - School of Medicine - Department of Biochemistry and Nutrition, ايران , Javadi, Maryam qazvin university of medical sciences - School of Medicine - Department of Nutrition and Dietary Therapy, ايران , Bandehpour, Mojgan shahid beheshti university of medical sciences - Cellular and Molecular Biology Research Center, School of Medicine - Department of Biotechnology, تهران, ايران , Karimi, Jamshid hamadan university of medical sciences - School of Medicine - Department of Biochemistry and Nutrition, ايران , Ziaee, Amir qazvin university of medical sciences - Qazvin Metabolic Disease Research Center - Department of Endocrinology, ايران , Koochaki, Amaneh shahid beheshti university of medical sciences - Cellular and Molecular Biology Research Center, School of Medicine - Department of Biotechnology, تهران, ايران , Torabi, Ali Pasteur Institute of Iran - Influenza Unit, ايران , Goodarzi, Mohammad Taghi hamadan university of medical sciences - Research Center for Molecular Medicine, ايران
Abstract :
Background: Diabetes mellitus type 1, formerly called insulin-dependent diabetes, is one of the autoimmune diseases where insulin-producing cells are destroyed by autoimmune response via T cells. The new approaches in treatment of diabetes are using the stem cells, cell transplantation of islet β cell, gene transfer by virus based plasmids, and non-viral gene constructs. Objectives: The purpose of this study was to construct glucose inducible insulin gene plasmid and use it in the muscle tissue of the rabbit. Materials and Methods: To achieve this goal, the preproinsulin, metallothionein2A promoter and the response element to carbohydrate genes were cloned into pBIND plasmid by standard cloning methods, to construct pBINDMTChIns. The gene cloning products were confirmed by the polymerase chain reaction (PCR) and restriction enzyme digestion template. The recombinant plasmid, containing the preproinsulin gene, was transferred into NIH3T3 cells and insulin gene expression was evaluated by reverse transcriptase PCR and western blotting techniques. Plasmid naked DNA containing the preproinsulin gene was injected into the rabbits’ thigh muscles, and its expression was confirmed by western blotting method. Results: This study shows the prepared gene construct is inducible by glucose. Gene expression of preproinsulin was observed in muscle tissue of rabbits. Conclusions: These finding indicated that research in diabetes mellitus gene therapy could be performed on larger animals.
Keywords :
Diabetes Mellitus , Preproinsulin , Rabbits , Gene Therapy
