A nonviral pHEMA+chitosan nanosphere-mediated high-efficiency gene delivery system

A nonviral pHEMA+chitosan nanosphere-mediated high-efficiency gene delivery system

Erdal Eroglu,1 Pooja M Tiwari,1 Alain B Waffo,1 Michael E Miller,2 Komal Vig,1 Vida A Dennis,1 Shree R Singh1 1Center for NanoBiotechnology Research, Alabama State University, Montgomery, AL, USA; 2Research Instrumentation Facility, Auburn University, AL, USA Abstract: The transport of DNA into euka...

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Autores principales: Eroglu E, Tiwari PM, Waffo AB, Miller ME, Vig K, Dennis VA, Singh SR
Formato: article
Lenguaje:EN
Publicado: Dove Medical Press 2013
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Medicine (General)
R5-920
Acceso en línea:https://doaj.org/article/89c54a5ee936450893ee2e64a0a13dae
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spelling oai:doaj.org-article:89c54a5ee936450893ee2e64a0a13dae2021-12-02T00:21:18ZA nonviral pHEMA+chitosan nanosphere-mediated high-efficiency gene delivery system1176-91141178-2013https://doaj.org/article/89c54a5ee936450893ee2e64a0a13dae2013-04-01T00:00:00Zhttp://www.dovepress.com/a-nonviral-phemachitosan-nanosphere-mediated-high-efficiency-gene-deli-a12726https://doaj.org/toc/1176-9114https://doaj.org/toc/1178-2013Erdal Eroglu,1 Pooja M Tiwari,1 Alain B Waffo,1 Michael E Miller,2 Komal Vig,1 Vida A Dennis,1 Shree R Singh1 1Center for NanoBiotechnology Research, Alabama State University, Montgomery, AL, USA; 2Research Instrumentation Facility, Auburn University, AL, USA Abstract: The transport of DNA into eukaryotic cells is minimal because of the cell membrane barrier, and this limits the application of DNA vaccines, gene silencing, and gene therapy. Several available transfection reagents and techniques have been used to circumvent this problem. Alternatively, nonviral nanoscale vectors have been shown to bypass the eukaryotic cell membrane. In the present work, we developed a unique nanomaterial, pHEMA+chitosan nanospheres (PCNSs), which consisted of poly (2-hydroxyethyl methacrylate) nanospheres surrounded by a chitosan cationic shell, and we used this for encapsulation of a respiratory syncytial virus (RSV)-F gene construct (a model for a DNA vaccine). The new nanomaterial was capable of transfecting various eukaryotic cell lines without the use of a commercial transfection reagent. Using transmission electron microscopy, (TEM), fluorescence activated cell sorting (FACS), and immunofluorescence, we clearly demonstrated that the positively charged PCNSs were able to bind to the negatively charged cell membrane and were taken up by endocytosis, in Cos-7 cells. Using quantitative polymerase chain reaction (qPCR), we also evaluated the efficiency of transfection achieved with PCNSs and without the use of a liposomal-based transfection mediator, in Cos-7, HEp-2, and Vero cells. To assess the transfection efficiency of the PCNSs in vivo, these novel nanomaterials containing RSV-F gene were injected intramuscularly into BALB/c mice, resulting in high copy number of the transgene. In this study, we report, for the first time, the application of the PCNSs as a nanovehicle for gene delivery in vitro and in vivo. Keywords: pHEMA+chitosan nanoparticles, nonviral vector, RSV-DNA vaccineEroglu ETiwari PMWaffo ABMiller MEVig KDennis VASingh SRDove Medical PressarticleMedicine (General)R5-920ENInternational Journal of Nanomedicine, Vol 2013, Iss default, Pp 1403-1415 (2013)
institution DOAJ
collection DOAJ
language EN
topic Medicine (General)
R5-920
spellingShingle Medicine (General)
R5-920
Eroglu E
Tiwari PM
Waffo AB
Miller ME
Vig K
Dennis VA
Singh SR
A nonviral pHEMA+chitosan nanosphere-mediated high-efficiency gene delivery system
description Erdal Eroglu,1 Pooja M Tiwari,1 Alain B Waffo,1 Michael E Miller,2 Komal Vig,1 Vida A Dennis,1 Shree R Singh1 1Center for NanoBiotechnology Research, Alabama State University, Montgomery, AL, USA; 2Research Instrumentation Facility, Auburn University, AL, USA Abstract: The transport of DNA into eukaryotic cells is minimal because of the cell membrane barrier, and this limits the application of DNA vaccines, gene silencing, and gene therapy. Several available transfection reagents and techniques have been used to circumvent this problem. Alternatively, nonviral nanoscale vectors have been shown to bypass the eukaryotic cell membrane. In the present work, we developed a unique nanomaterial, pHEMA+chitosan nanospheres (PCNSs), which consisted of poly (2-hydroxyethyl methacrylate) nanospheres surrounded by a chitosan cationic shell, and we used this for encapsulation of a respiratory syncytial virus (RSV)-F gene construct (a model for a DNA vaccine). The new nanomaterial was capable of transfecting various eukaryotic cell lines without the use of a commercial transfection reagent. Using transmission electron microscopy, (TEM), fluorescence activated cell sorting (FACS), and immunofluorescence, we clearly demonstrated that the positively charged PCNSs were able to bind to the negatively charged cell membrane and were taken up by endocytosis, in Cos-7 cells. Using quantitative polymerase chain reaction (qPCR), we also evaluated the efficiency of transfection achieved with PCNSs and without the use of a liposomal-based transfection mediator, in Cos-7, HEp-2, and Vero cells. To assess the transfection efficiency of the PCNSs in vivo, these novel nanomaterials containing RSV-F gene were injected intramuscularly into BALB/c mice, resulting in high copy number of the transgene. In this study, we report, for the first time, the application of the PCNSs as a nanovehicle for gene delivery in vitro and in vivo. Keywords: pHEMA+chitosan nanoparticles, nonviral vector, RSV-DNA vaccine
format article
author Eroglu E
Tiwari PM
Waffo AB
Miller ME
Vig K
Dennis VA
Singh SR
author_facet Eroglu E
Tiwari PM
Waffo AB
Miller ME
Vig K
Dennis VA
Singh SR
author_sort Eroglu E
title A nonviral pHEMA+chitosan nanosphere-mediated high-efficiency gene delivery system
title_short A nonviral pHEMA+chitosan nanosphere-mediated high-efficiency gene delivery system
title_full A nonviral pHEMA+chitosan nanosphere-mediated high-efficiency gene delivery system
title_fullStr A nonviral pHEMA+chitosan nanosphere-mediated high-efficiency gene delivery system
title_full_unstemmed A nonviral pHEMA+chitosan nanosphere-mediated high-efficiency gene delivery system
title_sort nonviral phema+chitosan nanosphere-mediated high-efficiency gene delivery system
publisher Dove Medical Press
publishDate 2013
url https://doaj.org/article/89c54a5ee936450893ee2e64a0a13dae
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