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Activation of pluripotency-associated genes in mouse embryonic fibroblasts by non-viral transfection with in vitro-derived mRNAs encoding Oct4, Sox2, Klf4 and cMyc

Paper ID Volume ID Publish Year Pages File Format Full-Text
7607 558 2012 6 PDF Available
Title
Activation of pluripotency-associated genes in mouse embryonic fibroblasts by non-viral transfection with in vitro-derived mRNAs encoding Oct4, Sox2, Klf4 and cMyc
Abstract

The first successful reprogramming of differentiated cells to a pluripotent state was done by retroviral introduction of four transcription factors (Oct4, Sox2, Klf4, cMyc) by the group of Yamanaka in 2006. Since then, scientists all over the world have attempted various methods to avoid insertional mutagenesis, a major limitation of the retrovirus-based method, however no technique was found to completely avoid DNA integration. Recently, a non-viral mRNA-based approach, inherent to avoid genomic integration, was implemented to generate stem cell-like cells, yet, seventeen daily transfections were required, inducing substantial stress on the cells. In this work, we demonstrate successful activation of pluripotency-associated genes in mouse embryonic fibroblasts by means of cationic lipid-mediated introduction of mRNAs encoding the four factors. Moreover, our transfection protocol required maximally three transfections. Up-regulation of the transfected factors as well as Nanog and SSEA-1, typical mouse pluripotency markers, was detected already after the first transfection. Nuclear localization of the introduced factors was confirmed. Positive alkaline phosphatase staining of cell clusters further confirmed the onset of the reprogramming process. In conclusion, the transfection method presented here holds great promise for safe generation of induced pluripotent stem cells of mouse origin.

Keywords
mRNA; Reprogramming; Non-viral; Liposome; Oct4
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Activation of pluripotency-associated genes in mouse embryonic fibroblasts by non-viral transfection with in vitro-derived mRNAs encoding Oct4, Sox2, Klf4 and cMyc
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Publisher
Database: Elsevier - ScienceDirect
Journal: Biomaterials - Volume 33, Issue 2, January 2012, Pages 412–417
Authors
, , , , , ,
Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us