Efficient TRAIL Gene Delivery Using Nucleofection Based Method in Human Adiposed Derived Mesenchymal Stromal Cells

Kamal Shaik Fakiruddin; Puteri Baharuddin; Moon Nian Lim; Noor Atiqah Fakharuzi; Nurul Ain Nasim; Zubaidah Zakaria

doi:doi:10.11648/j.cb.20140201.11

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Efficient TRAIL Gene Delivery Using Nucleofection Based Method in Human Adiposed Derived Mesenchymal Stromal Cells

Kamal Shaik Fakiruddin, Puteri Baharuddin, Moon Nian Lim, Noor Atiqah Fakharuzi, Nurul Ain Nasim, Zubaidah Zakaria

Published in Cell Biology (Volume 2, Issue 1)

Received: 15 May 2014 Accepted: 23 May 2014 Published: 10 June 2014

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Abstract

Being able to perform genetic manipulation of human adipose-derived mesenchymal stromal cells (ADMSCs) will harness the benefits of these cells beyond degenerative diseases. Most primary cells show resistance to genetic alteration with viral transduction remains to be the most effective tool for gene delivery. However, the use of viral vectors has several disadvantages mainly involving safety risk. Here, we report optimization using safe and yet efficient nucleofection based transfection of DNA plasmid encoded for TNF-related apoptosis inducing ligand (TRAIL) into ADMSCs. Initial characterization of ADMSCs was performed based on cells morphological evaluation and surface protein expression. Nucleofection revealed 10% higher transfection efficiency compared to lipofection (Fugene 6 and Turbofect) with optimal cells viability (~87%). Subsequent nucleofection analysis showed the increased plasmid concentration of 10µg resulted in significantly higher reporter expression with 35% efficiency and 43% yield. Transgene expression was stable at day 9 with 74% cells remained to be GFP+, but was reduced to baseline at day 15. In this report, we have showed that the nucleofection technique is efficient to deliver exogenous gene in ADMSCs compared to common lipofection methods. We also noticed that increased plasmid concentration enhanced nucleofection efficiency and yield in ADMSC. Furthermore, exogenous expression of the gene was transient with no evidence of stable genomic integration, thus we concluded that the nucleofection technique is an efficient and yet safe nonviral transfection technique in ADMSCs.

Published in	Cell Biology (Volume 2, Issue 1)
DOI	10.11648/j.cb.20140201.11
Page(s)	1-6
Creative Commons	This is an Open Access article, distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution and reproduction in any medium or format, provided the original work is properly cited.
Copyright	Copyright © The Author(s), 2014. Published by Science Publishing Group

Keywords

Human Adiposed Derived Mesenchymal Stromal Cells, TRAIL, Gene Transfection, Nucleofection

References

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[7]	Almasan, Alexandru and Ashkenazi, Avi, “Apo2L/TRAIL: apoptosis signalling, biology, and potential for cancer therapy,” Cytokine & Growth Factor Reviews, vol.14 (3-4), pp.337-48, 2003.
[8]	Gresch, Oliver, et al., “New non-viral method for gene transfer into primary cells,” Methods, vol.33 (2), pp.151-63, 2004.
[9]	Cesnulevicius, Konstantin, et al., “Nucleofection Is the Most Efficient Nonviral Transfection Method for Neuronal Stem Cells Derived from Ventral Mesencephali with No Changes in Cell Composition or Dopaminergic Fate,” STEM CELLS, vol.24 (12), pp.2776-91, 2006.
[10]	Distler, Jörg H. W., et al., “Nucleofection: a new, highly efficient transfection method for primary human keratinocytes,” Experimental Dermatology, vol.14 (4), pp.315-20, 2005.
[11]	Bartholomew, Amelia, et al., “Mesenchymal stem cells suppress lym-phocyte proliferation in vitro and prolong skin graft survival in vivo,” Experimental Hematology, vol.30 (1), pp.42-48, 2002.
[12]	Le Blanc, Katarina, et al., “Treatment of severe acute graft-versus-host disease with third party haploidentical mesenchymal stem cells,” The Lancet, vol.363 (9419), pp.1439-41, 2004.
[13]	Amit N Patel, Jorge Genovese, “Potential clinical ap-plications of adult human mesenchymal stem cell (Prochymal®) therapy,” Stem Cells and Cloning: Advances and Applications, vol.4, pp.61-72, 2011.
[14]	Lu, Yan-rong, et al., “The growth inhi-bitory effect of mesenchymal stem cells on tumor cells in vitro and in vivo,” Cancer Biology & Therapy, vol.7 (2), pp. 245-51, 2008.
[15]	Nakamura, K., et al. (2004), 'Antitumor effect of ge-netically engineered mesenchymal stem cells in a rat glioma model,” Gene Ther, 11 (14), 1155-64.
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Kamal Shaik Fakiruddin, Puteri Baharuddin, Moon Nian Lim, Noor Atiqah Fakharuzi, Nurul Ain Nasim, et al. (2014). Efficient TRAIL Gene Delivery Using Nucleofection Based Method in Human Adiposed Derived Mesenchymal Stromal Cells. Cell Biology, 2(1), 1-6. https://doi.org/10.11648/j.cb.20140201.11

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ACS Style

Kamal Shaik Fakiruddin; Puteri Baharuddin; Moon Nian Lim; Noor Atiqah Fakharuzi; Nurul Ain Nasim, et al. Efficient TRAIL Gene Delivery Using Nucleofection Based Method in Human Adiposed Derived Mesenchymal Stromal Cells. Cell Biol. 2014, 2(1), 1-6. doi: 10.11648/j.cb.20140201.11

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AMA Style

Kamal Shaik Fakiruddin, Puteri Baharuddin, Moon Nian Lim, Noor Atiqah Fakharuzi, Nurul Ain Nasim, et al. Efficient TRAIL Gene Delivery Using Nucleofection Based Method in Human Adiposed Derived Mesenchymal Stromal Cells. Cell Biol. 2014;2(1):1-6. doi: 10.11648/j.cb.20140201.11

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@article{10.11648/j.cb.20140201.11,
  author = {Kamal Shaik Fakiruddin and Puteri Baharuddin and Moon Nian Lim and Noor Atiqah Fakharuzi and Nurul Ain Nasim and Zubaidah Zakaria},
  title = {Efficient TRAIL Gene Delivery Using Nucleofection Based Method in Human Adiposed Derived Mesenchymal Stromal Cells},
  journal = {Cell Biology},
  volume = {2},
  number = {1},
  pages = {1-6},
  doi = {10.11648/j.cb.20140201.11},
  url = {https://doi.org/10.11648/j.cb.20140201.11},
  eprint = {https://article.sciencepublishinggroup.com/pdf/10.11648.j.cb.20140201.11},
  abstract = {Being able to perform genetic manipulation of human adipose-derived mesenchymal stromal cells (ADMSCs) will harness the benefits of these cells beyond degenerative diseases. Most primary cells show resistance to genetic alteration with viral transduction remains to be the most effective tool for gene delivery. However, the use of viral vectors has several disadvantages mainly involving safety risk. Here, we report optimization using safe and yet efficient nucleofection based transfection of DNA plasmid encoded for TNF-related apoptosis inducing ligand (TRAIL) into ADMSCs. Initial characterization of ADMSCs was performed based on cells morphological evaluation and surface protein expression. Nucleofection revealed 10% higher transfection efficiency compared to lipofection (Fugene 6 and Turbofect) with optimal cells viability (~87%). Subsequent nucleofection analysis showed the increased plasmid concentration of 10µg resulted in significantly higher reporter expression with 35% efficiency and 43% yield. Transgene expression was stable at day 9 with 74% cells remained to be GFP+, but was reduced to baseline at day 15. In this report, we have showed that the nucleofection technique is efficient to deliver exogenous gene in ADMSCs compared to common lipofection methods. We also noticed that increased plasmid concentration enhanced nucleofection efficiency and yield in ADMSC. Furthermore, exogenous expression of the gene was transient with no evidence of stable genomic integration, thus we concluded that the nucleofection technique is an efficient and yet safe nonviral transfection technique in ADMSCs.},
 year = {2014}
}

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TY  - JOUR
T1  - Efficient TRAIL Gene Delivery Using Nucleofection Based Method in Human Adiposed Derived Mesenchymal Stromal Cells
AU  - Kamal Shaik Fakiruddin
AU  - Puteri Baharuddin
AU  - Moon Nian Lim
AU  - Noor Atiqah Fakharuzi
AU  - Nurul Ain Nasim
AU  - Zubaidah Zakaria
Y1  - 2014/06/10
PY  - 2014
N1  - https://doi.org/10.11648/j.cb.20140201.11
DO  - 10.11648/j.cb.20140201.11
T2  - Cell Biology
JF  - Cell Biology
JO  - Cell Biology
SP  - 1
EP  - 6
PB  - Science Publishing Group
SN  - 2330-0183
UR  - https://doi.org/10.11648/j.cb.20140201.11
AB  - Being able to perform genetic manipulation of human adipose-derived mesenchymal stromal cells (ADMSCs) will harness the benefits of these cells beyond degenerative diseases. Most primary cells show resistance to genetic alteration with viral transduction remains to be the most effective tool for gene delivery. However, the use of viral vectors has several disadvantages mainly involving safety risk. Here, we report optimization using safe and yet efficient nucleofection based transfection of DNA plasmid encoded for TNF-related apoptosis inducing ligand (TRAIL) into ADMSCs. Initial characterization of ADMSCs was performed based on cells morphological evaluation and surface protein expression. Nucleofection revealed 10% higher transfection efficiency compared to lipofection (Fugene 6 and Turbofect) with optimal cells viability (~87%). Subsequent nucleofection analysis showed the increased plasmid concentration of 10µg resulted in significantly higher reporter expression with 35% efficiency and 43% yield. Transgene expression was stable at day 9 with 74% cells remained to be GFP+, but was reduced to baseline at day 15. In this report, we have showed that the nucleofection technique is efficient to deliver exogenous gene in ADMSCs compared to common lipofection methods. We also noticed that increased plasmid concentration enhanced nucleofection efficiency and yield in ADMSC. Furthermore, exogenous expression of the gene was transient with no evidence of stable genomic integration, thus we concluded that the nucleofection technique is an efficient and yet safe nonviral transfection technique in ADMSCs.
VL  - 2
IS  - 1
ER  -

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Author Information

Kamal Shaik Fakiruddin

Stem Cell Laboratory, Cancer Research Centre, Institute for Medical Research (IMR), Kuala Lumpur Malaysia
Puteri Baharuddin

Stem Cell Laboratory, Cancer Research Centre, Institute for Medical Research (IMR), Kuala Lumpur Malaysia
Moon Nian Lim

Stem Cell Laboratory, Cancer Research Centre, Institute for Medical Research (IMR), Kuala Lumpur Malaysia
Noor Atiqah Fakharuzi

Stem Cell Laboratory, Cancer Research Centre, Institute for Medical Research (IMR), Kuala Lumpur Malaysia
Nurul Ain Nasim

Stem Cell Laboratory, Cancer Research Centre, Institute for Medical Research (IMR), Kuala Lumpur Malaysia
Zubaidah Zakaria

Stem Cell Laboratory, Cancer Research Centre, Institute for Medical Research (IMR), Kuala Lumpur Malaysia

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APA Style

Kamal Shaik Fakiruddin, Puteri Baharuddin, Moon Nian Lim, Noor Atiqah Fakharuzi, Nurul Ain Nasim, et al. (2014). Efficient TRAIL Gene Delivery Using Nucleofection Based Method in Human Adiposed Derived Mesenchymal Stromal Cells. Cell Biology, 2(1), 1-6. https://doi.org/10.11648/j.cb.20140201.11

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ACS Style

Kamal Shaik Fakiruddin; Puteri Baharuddin; Moon Nian Lim; Noor Atiqah Fakharuzi; Nurul Ain Nasim, et al. Efficient TRAIL Gene Delivery Using Nucleofection Based Method in Human Adiposed Derived Mesenchymal Stromal Cells. Cell Biol. 2014, 2(1), 1-6. doi: 10.11648/j.cb.20140201.11

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AMA Style

Kamal Shaik Fakiruddin, Puteri Baharuddin, Moon Nian Lim, Noor Atiqah Fakharuzi, Nurul Ain Nasim, et al. Efficient TRAIL Gene Delivery Using Nucleofection Based Method in Human Adiposed Derived Mesenchymal Stromal Cells. Cell Biol. 2014;2(1):1-6. doi: 10.11648/j.cb.20140201.11

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@article{10.11648/j.cb.20140201.11,
  author = {Kamal Shaik Fakiruddin and Puteri Baharuddin and Moon Nian Lim and Noor Atiqah Fakharuzi and Nurul Ain Nasim and Zubaidah Zakaria},
  title = {Efficient TRAIL Gene Delivery Using Nucleofection Based Method in Human Adiposed Derived Mesenchymal Stromal Cells},
  journal = {Cell Biology},
  volume = {2},
  number = {1},
  pages = {1-6},
  doi = {10.11648/j.cb.20140201.11},
  url = {https://doi.org/10.11648/j.cb.20140201.11},
  eprint = {https://article.sciencepublishinggroup.com/pdf/10.11648.j.cb.20140201.11},
  abstract = {Being able to perform genetic manipulation of human adipose-derived mesenchymal stromal cells (ADMSCs) will harness the benefits of these cells beyond degenerative diseases. Most primary cells show resistance to genetic alteration with viral transduction remains to be the most effective tool for gene delivery. However, the use of viral vectors has several disadvantages mainly involving safety risk. Here, we report optimization using safe and yet efficient nucleofection based transfection of DNA plasmid encoded for TNF-related apoptosis inducing ligand (TRAIL) into ADMSCs. Initial characterization of ADMSCs was performed based on cells morphological evaluation and surface protein expression. Nucleofection revealed 10% higher transfection efficiency compared to lipofection (Fugene 6 and Turbofect) with optimal cells viability (~87%). Subsequent nucleofection analysis showed the increased plasmid concentration of 10µg resulted in significantly higher reporter expression with 35% efficiency and 43% yield. Transgene expression was stable at day 9 with 74% cells remained to be GFP+, but was reduced to baseline at day 15. In this report, we have showed that the nucleofection technique is efficient to deliver exogenous gene in ADMSCs compared to common lipofection methods. We also noticed that increased plasmid concentration enhanced nucleofection efficiency and yield in ADMSC. Furthermore, exogenous expression of the gene was transient with no evidence of stable genomic integration, thus we concluded that the nucleofection technique is an efficient and yet safe nonviral transfection technique in ADMSCs.},
 year = {2014}
}

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TY  - JOUR
T1  - Efficient TRAIL Gene Delivery Using Nucleofection Based Method in Human Adiposed Derived Mesenchymal Stromal Cells
AU  - Kamal Shaik Fakiruddin
AU  - Puteri Baharuddin
AU  - Moon Nian Lim
AU  - Noor Atiqah Fakharuzi
AU  - Nurul Ain Nasim
AU  - Zubaidah Zakaria
Y1  - 2014/06/10
PY  - 2014
N1  - https://doi.org/10.11648/j.cb.20140201.11
DO  - 10.11648/j.cb.20140201.11
T2  - Cell Biology
JF  - Cell Biology
JO  - Cell Biology
SP  - 1
EP  - 6
PB  - Science Publishing Group
SN  - 2330-0183
UR  - https://doi.org/10.11648/j.cb.20140201.11
AB  - Being able to perform genetic manipulation of human adipose-derived mesenchymal stromal cells (ADMSCs) will harness the benefits of these cells beyond degenerative diseases. Most primary cells show resistance to genetic alteration with viral transduction remains to be the most effective tool for gene delivery. However, the use of viral vectors has several disadvantages mainly involving safety risk. Here, we report optimization using safe and yet efficient nucleofection based transfection of DNA plasmid encoded for TNF-related apoptosis inducing ligand (TRAIL) into ADMSCs. Initial characterization of ADMSCs was performed based on cells morphological evaluation and surface protein expression. Nucleofection revealed 10% higher transfection efficiency compared to lipofection (Fugene 6 and Turbofect) with optimal cells viability (~87%). Subsequent nucleofection analysis showed the increased plasmid concentration of 10µg resulted in significantly higher reporter expression with 35% efficiency and 43% yield. Transgene expression was stable at day 9 with 74% cells remained to be GFP+, but was reduced to baseline at day 15. In this report, we have showed that the nucleofection technique is efficient to deliver exogenous gene in ADMSCs compared to common lipofection methods. We also noticed that increased plasmid concentration enhanced nucleofection efficiency and yield in ADMSC. Furthermore, exogenous expression of the gene was transient with no evidence of stable genomic integration, thus we concluded that the nucleofection technique is an efficient and yet safe nonviral transfection technique in ADMSCs.
VL  - 2
IS  - 1
ER  -

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