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Stem Cells International Volume 2019 ,2019-01-17
Comparative AAV-eGFP Transgene Expression Using Vector Serotypes 1–9, 7m8, and 8b in Human Pluripotent Stem Cells, RPEs, and Human and Rat Cortical Neurons
Research Article
Thu T. Duong 1 James Lim 2 Vidyullatha Vasireddy 1 Tyler Papp 1 Hung Nguyen 3 Lanfranco Leo 1 Jieyan Pan 1 Shangzhen Zhou 1 H. Isaac Chen 2 , 4 Jean Bennett 1 Jason A. Mills 1
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Received 2018-04-23, accepted for publication 2018-11-16, Published 2018-11-16

Recombinant adeno-associated virus (rAAV), produced from a nonpathogenic parvovirus, has become an increasing popular vector for gene therapy applications in human clinical trials. However, transduction and transgene expression of rAAVs can differ across in vitro and ex vivo cellular transduction strategies. This study compared 11 rAAV serotypes, carrying one reporter transgene cassette containing a cytomegalovirus immediate-early enhancer (eCMV) and chicken beta actin (CBA) promoter driving the expression of an enhanced green-fluorescent protein (eGFP) gene, which was transduced into four different cell types: human iPSC, iPSC-derived RPE, iPSC-derived cortical, and dissociated embryonic day 18 rat cortical neurons. Each cell type was exposed to three multiplicity of infections (MOI: 1E4, 1E5, and 1E6 vg/cell). After 24, 48, 72, and 96 h posttransduction, GFP-expressing cells were examined and compared across dosage, time, and cell type. Retinal pigmented epithelium showed highest AAV-eGFP expression and iPSC cortical the lowest. At an MOI of 1E6 vg/cell, all serotypes show measurable levels of AAV-eGFP expression; moreover, AAV7m8 and AAV6 perform best across MOI and cell type. We conclude that serotype tropism is not only capsid dependent but also cell type plays a significant role in transgene expression dynamics.


Copyright © 2019 Thu T. Duong et al. 2019
This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


Jason A. Mills.F.M. Kirby Center for Molecular Ophthalmology and Center for Advanced Retinal and Ocular Therapeutics (CAROT), Scheie Eye Institute, University of Pennsylvania Perelman School of Medicine, Pennsylvania, PA 19104, USA, upenn.edu.millsja@pennmedicine.upenn.edu


Thu T. Duong,James Lim,Vidyullatha Vasireddy,Tyler Papp,Hung Nguyen,Lanfranco Leo,Jieyan Pan,Shangzhen Zhou,H. Isaac Chen,Jean Bennett,Jason A. Mills. Comparative AAV-eGFP Transgene Expression Using Vector Serotypes 1–9, 7m8, and 8b in Human Pluripotent Stem Cells, RPEs, and Human and Rat Cortical Neurons. Stem Cells International ,Vol.2019(2019)



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