Efficient TRAIL Gene Delivery Using Nucleofection Based Method in Human Adiposed Derived Mesenchymal Stromal Cells
Cell Biology
Volume 2, Issue 1, January 2014, Pages: 1-6
Received: May 15, 2014; Accepted: May 23, 2014; Published: Jun. 10, 2014
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Authors
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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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.
Keywords
Human Adiposed Derived Mesenchymal Stromal Cells, TRAIL, Gene Transfection, Nucleofection
To cite this article
Kamal Shaik Fakiruddin, Puteri Baharuddin, Moon Nian Lim, Noor Atiqah Fakharuzi, Nurul Ain Nasim, Zubaidah Zakaria, Efficient TRAIL Gene Delivery Using Nucleofection Based Method in Human Adiposed Derived Mesenchymal Stromal Cells, Cell Biology. Vol. 2, No. 1, 2014, pp. 1-6. doi: 10.11648/j.cb.20140201.11
References
[1]
Barry, Frank P. and Murphy, J. Mary, “Mesenchymal stem cells: clinical applications and biological characterization,” The International Journal of Biochemistry & Cell Biology, vol.36 (4), pp.568-84, 2004.
[2]
Dominici, M., et al. “Minimal criteria for defining multipotent mesenchymal stromal cells,” The International Society for Cellular Therapy position statement', Cytotherapy, vol.8 (4), pp.315-17, 2006.
[3]
Le Blanc, Katarina, et al., “HLA expression and immunologic propertie-sof differentiated and undifferentiated mesenchymal stem cells,” Experimental Hematology, vol.31 (10), pp.890-96, 2003.
[4]
Rodriguez, A. M., et al. “The human adipose tissue is a source of multipotent stem cells,” Biochimie, vol.87 (1), pp.125-28, 2005.
[5]
Stagg, John, “Mesenchymal Stem Cells in Cancer,” Stem Cell Reviews, vol.4 (2), pp.119-24, 2008.
[6]
Guillot, Pascale V., et al., “Stem cell differentiation and expansion for clinical appli-cations of tissue engineering,” Journal of Cellular and Molecular Medicine, vol.11 (5), pp.935-44, 2007.
[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.
[16]
Hana Haleem-Smith, Assia Derfoul, Chukwuka Okafor, Richard Tuli, Douglas Ol-sen, David J. Hall, Rocky S. Tuan, “Optimization of High Efficiency Transfection of Adult Human Mesenchymal stem Cells In Vitro,” Molecular Biotechnology, vol.30, pp.9-19, 2005 .
[17]
Bellail, A. C., et al., “TRAIL agonists on clinical trials for cancer therapy: the promises and the challenges,” Rev Recent Clin Trials, vol.4 (1), pp.34-41, 2009.
[18]
Choi, Seung Ah, et al., “Therapeutic efficacy and safety of TRAIL-producing human adipose tissue–derived mesenchymal stem cells against experimental brainstem glioma,” Neuro-Oncology, vol.13 (1), pp.61-69, 2011.
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