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S. Shahrokhi* 1 , K. Alimoghaddam 2 , A. Ghavamzadeh 2

Role of Substance P (SP) and Calcitonin Gene-related Peptide (CGRP) in Gibbon-Ape-Leukemia Virus (GALV) Transduction of CD34 + Cells. S. Shahrokhi* 1 , K. Alimoghaddam 2 , A. Ghavamzadeh 2.

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S. Shahrokhi* 1 , K. Alimoghaddam 2 , A. Ghavamzadeh 2

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  1. Role of Substance P (SP) and Calcitonin Gene-related Peptide (CGRP)in Gibbon-Ape-Leukemia Virus (GALV) Transduction of CD34+ Cells S. Shahrokhi*1,K. Alimoghaddam2, A. Ghavamzadeh2 1 Department of Immunology, School of Medicine, Lorestan University of Medical Sciences, Khorramabad, Iran. *email:shahrokhi_so@yahoo.com 2Hematology, oncology and stem cell transplantation research center of Tehran University of Medical Sciences, Tehran, Iran. Results: Viral vectors titration on Hela cells indicated transduction efficiency of 1×106CFU/ml. Real Time PCR of Neo and CFU-C showed stimulatory role of SP on gene transfer 5.9 and 14.84% compared to 3.6 and 12.58% in control group, while opposite role observed for CGRP 0.89 and 7.86%. Both SP and CGRP showed no significant effect in these assays. Conclusions:This study showed including of SP in growth factor cocktail is beneficial for CD34+ transduction, which could be applied to genetic modification procedures. Introduction: Optimization of transduction condition is an important goal to improve gene transduction. Therefore, we aimed to assess the effect of SP and/or CGRP as novel growth/transducing factors on the efficacy of CD34+ transduction. . Material and Methods: CD34+ cells were transduced with Gibbon-Ape-Leukemia virus (GALV), containing Neomycin gene. CD34+ cells were transduced with GALV presence of SP and/or CGRP. Real Time PCR and colony formation assay (CFU-C) was performed. Figure 1. Evaluation of transduction efficacy via CFU-C in non/transduced CD34+ cells, in the presence and absence of SP and/or CGRP neuropeptides. Following CD34+ cells transduction, cells harvested and cultivated for extended 14 days in CFU-C assay. Efficacy of transduction was enumerated by the ratio of G418 resistance CFU-GM colony by total CFU- GM colony in G418 minus plate. No colony growths in Mock infected cells. * Represents a statistically significant (p <0.05) difference to control group. Figure 2.Quantification of Neo gene expression in non/transduced CD34+, in the presence and absence of SP and/or CGRP neuropeptides. Concentrations of 10-9M of SP and/ or CGRP were added in addition to the cytokine cocktail to isolated CD34+ cells. The control group was expanded in the cytokine cocktail alone. Following 24 hrs, GALV was added every 12 hrs for 4 times. DNA of neuropeptide-cytokine treated cells and the control group was extracted and the amount of Neo gene expression in non/transduced CD34+ cells was evaluated by Real Time PCR analysis. * Represents a statistically significant (p <0.05) difference to control group.

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