Cellular transfection with calcium phosphate/DNA nanoparticles – low intracellular calcium disturbance and further developments by PEI-functionalization

Sebastian Neumann¹, Viktoriya Sokolova², Anna Kovtun³, Svitlana Chernousova⁴, Irmgard D. Dietzel⁵, Matthias Epple⁶, Rolf Heumann⁷

1 Department of Biochemistry, Molecular Neurobiochemistry, Ruhr-University Bochum, 44780 Bochum, Germany

2 Institute of Inorganic Chemistry and Center for Nanointegration Duisburg-Essen (CeNIDE), University of Duisburg-Essen, 45117 Essen, Germany

3 Institute of Inorganic Chemistry and Center for Nanointegration Duisburg-Essen (CeNIDE), University of Duisburg-Essen, 45117 Essen, Germany

4 Institute of Inorganic Chemistry and Center for Nanointegration Duisburg-Essen (CeNIDE), University of Duisburg-Essen, 45117 Essen, Germany

5 Department of Biochemistry, Molecular Neurobiochemistry, Ruhr-University Bochum, 44780 Bochum, Germany

6 Institute of Inorganic Chemistry and Center for Nanointegration Duisburg-Essen (CeNIDE), University of Duisburg-Essen, 45117 Essen, Germany

7 Department of Biochemistry, Molecular Neurobiochemistry, Ruhr-University Bochum, 44780 Bochum, Germany

Abstract

For future application in the field of gene-therapy calcium phosphate nanoparticles can be functionalized with nucleic acids for its delivery into cells as recently reviewed in [1]. Using ⁴⁵calcium isotopic labelling and Fura-2 time-lapse fluorescence microscopy we analysed changes of the intracellular calcium level upon transfection with the standard calcium phosphate method versus calcium phosphate/DNA nanoparticles in the human bladder carcinoma cell line T24. Whereas the nanoparticles-mediated transfection slightly affected the intracellular calcium level the standard calcium phosphate method caused strong disturbances leading to cell death [2]. Furthermore the outer shell of the calcium phosphate/DNA nanoparticles was functionalized by poly(ethyleneimine) (PEI) yielding a positively charged surface. This increased the transfection efficiency in comparison with calcium phosphate/DNA nanoparticles without any PEI as shown in several cell lines (HeLa, T24 and NIH3T3). However, a compromise has to be found between the transfection efficiency and the amount of PEI due to its cytotoxicity [3].

Taken together, due to their good biocompatibility DNA-functionalised calcium phosphate nanoparticles may serve as a promising transfection agent for further in vivo studies.

References

[1] Epple, M., Ganesan, K., Heumann, R., Klesing, J., Kovtun, A., Neumann, S., and Sokolova, V. (2010). Application of calcium phosphate nanoparticles in biomedicine. JOURNAL OF MATERIALS CHEMISTRY 20, 18-23.

[2] Neumann, S., Kovtun, A., Dietzel, I.D., Epple, M., and Heumann, R. (2009). The use of size-defined DNA-functionalized calcium phosphate nanoparticles to minimise intracellular calcium disturbance during transfection. BIOMATERIALS 30, 6794-6802.

[3] Sokolova, V., Neumann, S., Kovtun, A., Chernousova, S., Heumann, R., and Epple, M. (2010). An outer shell of positively charged poly(ethyleneimine) strongly increases the transfection efficiency of calcium phosphate/DNA nanoparticles. JOURNAL OF MATERIALS SCIENCE 45, 4952-4957

DOI^®: 10.3288/contoo.paper.1494