Generation of hemophilia A mouse induced pluripotent stem cells using polycistronic lentiviral vector in serum- and feeder-free culture conditions

Abstract

Event Abstract Back to Event Generation of hemophilia A mouse induced pluripotent stem cells using polycistronic lentiviral vector in serum- and feeder-free culture conditions Rozita M. Sakri1, Syahril Abdullah2, 3 and Mohamed Saifulaman M. Said1* 1 Universiti Teknologi MARA, Faculty of Applied Sciences, Malaysia 2 Universiti Putra Malaysia, Genetics & Regenerative Medicine Research Centre, Faculty of Medicine & Health Sciences, Malaysia 3 Universiti Putra Malaysia, Institute of Bioscience, Malaysia Induced pluripotent stem cells (iPSCs) research has opened up exciting possibilities in many research areas especially in the field of regenerative medicine. The ability to reprogram back somatic cells into embryonic stem (ES)-like state has made patient-specific cells therapy possible unhindered ethical and practical dilemmas associated with the use of embryonic stem cells [1]. There have been several reports on the generation of iPS cells from murine somatic cells in vitro, however, there are still many challenges to overcome; (i) the induction efficiency is extremely low, (ii) multiple transgene integrations which increase the risk of tumorigenicity of iPS cells produced, (iii) partially differentiated iPS colonies due to incomplete reprogramming, (iv) the use of feeder cells and medium containing serum add complexity and variability in nutrients and factors that contribute to cell growth and the maintenance of pluripotency [2,3,4,5]. To overcome these issues, we utilized polycistronic lentiviral vector carrying the Yamanaka’s factors (Oct4, Sox2, Klf4 and c-Myc) for reprogramming of murine fibroblasts in serum- and feeder- free defined culture conditions. In this study, primary fibroblasts from hemophilic A B6;129S4-F8tm1Kaz/J and wild-type C57BL/6 mouse were transduced with an optimal multiplicity of infection of the virus produced. Suspected iPS cell colonies were picked within 20 days post-transduction. The results showed that iPS cells derived from the wild-type mouse fibroblast were successfully generated in serum- and feeder-free defined culture conditions. The generated wild-type mouse iPS cells are similar to embryonic stem cells in many aspects including morphology, their properties of self-renewal and pluripotency, and in vitro differentiation into the three primary germ layers. However, we were not able to generate iPS cells from hemophilia A mouse fibroblast with the similar lentiviral vector using the same reprogramming manner. We obtained neuronal-like morphology instead of the expected pluripotent cell colonies. The emerging of neuronal-like cells post-transduction is speculated as an incomplete reprogramming process [6]. These partially reprogrammed cells could not maintain the stem cells-like characteristic and eventually undergo differentiation. Hence, these data could provide an insight of reprogramming process. Acknowledgements Special thanks to Faculty of Applied Sciences, UiTM and Genetics & Regenerative Medicine Research Centre, UPM, for their assistance. This study was supported by ERGS Grant, MOHE. Keywords: induced pluripotent stem cell, cellular reprogramming, polycistronic lentivirus Conference: 6th Malaysian Tissue Engineering and Regenerative Medicine Scientific Meeting (6th MTERMS) 2016 and 2nd Malaysian Stem Cell Meeting, Seberang Jaya, Penang, Malaysia, 17 Nov - 18 Nov, 2016. Presentation Type: Poster Topic: Gene Therapy, Reprogramming and Pluripotency Citation: Sakri RM, Abdullah S and Said MM (2016). Generation of hemophilia A mouse induced pluripotent stem cells using polycistronic lentiviral vector in serum- and feeder-free culture conditions. Front. Bioeng. Biotechnol. Conference Abstract: 6th Malaysian Tissue Engineering and Regenerative Medicine Scientific Meeting (6th MTERMS) 2016 and 2nd Malaysian Stem Cell Meeting. doi: 10.3389/conf.FBIOE.2016.02.00028 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 08 Dec 2016; Published Online: 19 Dec 2016. * Correspondence: Dr. Mohamed Saifulaman M Said, Universiti Teknologi MARA, Faculty of Applied Sciences, Shah Alam, Selangor, 40450, Malaysia, dsaifulaman@salam.uitm.edu.my Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Rozita M Sakri Syahril Abdullah Mohamed Saifulaman M Said Google Rozita M Sakri Syahril Abdullah Mohamed Saifulaman M Said Google Scholar Rozita M Sakri Syahril Abdullah Mohamed Saifulaman M Said PubMed Rozita M Sakri Syahril Abdullah Mohamed Saifulaman M Said Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.