Projects / Programmes
Correlation of induced transmembrane voltage and electroporation-mediated transport of molecules on irregular shaped cells and cell clusters in vitro.
| Code |
Science |
Field |
Subfield |
| 2.06.07 |
Engineering sciences and technologies |
Systems and cybernetics |
Biomedical technics |
| Code |
Science |
Field |
| T115 |
Technological sciences |
Medical technology |
induced transmembrane voltage, electroporation, numerical modeling, finite elements method
Researchers (1)
| no. |
Code |
Name and surname |
Research area |
Role |
Period |
No. of publicationsNo. of publications |
| 1. |
22487 |
PhD Gorazd Pucihar |
Systems and cybernetics |
Head |
2007 - 2008 |
69 |
Organisations (1)
Abstract
When a biological cell is exposed to an external electric field, induced transmembrane voltage (ITV) forms on its membrane. When ITV exceeds some threshold value, the permeability of the cell membrane in these regions transiently increases. This phenomenon is termed electroporation. In many applications of electroporation an efficient and at the same time reversible electroporation is essential (e.g. DNA electrotransfer). Thus, a careful planning of the experiment, which involves the estimation of the amplitude of ITV leading to cell electroporation, is required. The problem arises in case of tissues, where cell geometry is more complicated, cells are close enough to affect the electric field around each other, and they are often connected with pathways between them. In all these cases, an analytical description of ITV is in general not attainable and numerical methods are often the only feasible approach. Due to the complexity of tissue structure, numerical models are either macroscopic, where detailed cell structure is not considered, or in case of microscopic models, they are constructed using simple geometrical shapes and do not reflect realistic tissue structure. Irregularly shaped cells and clusters of such cells are close to tissues in their complexity, at least regarding the shape, density and connections between cells, and could therefore represent considerably more realistic models of tissue than the commonly used simplified models. In the proposed research project we will first develop a method that will allow to construct realistic models of cells of irregular shapes and clusters of such cells. Then, the ITV, calculated on these models, will be compared with the ITV measured on the same cells and clusters used for model construction, and also with the electroporation-mediated transport of molecules, again on the same cells. Such calculations and measurements performed on these more realistic models of cells in tissue will provide a better understanding of electric field interactions with tissues on a microscopic (single cell) level, which in turn determines the macroscopic behavior of the tissue during electroporation. The findings of this research will improve our understanding of the course of electroporation in vivo, thereby providing the possibility to increase its efficiency, which is important in applications such as electrochemotherapy and electrogenetherapy.
Significance for science
The calculations and measurements of the induced transmembrane voltage (ITV), as well as monitoring of electroporation-mediated transport of molecules in cells of irregular shapes and clusters of such cells, present an original approach to studying the background and discovering the basic scientific principles of electroporation. More specifically, we discovered the following basic principles of electroporation:
(i) Cell shape, orientation, cell proximity and intracellular connections all have a profound influence on the ITV and electroporation and are responsible for the observed differences between the efficiency of electroporation in vitro and in vivo.
(ii) The ITV and electroporation mediated transport of molecules are correlated. Namely, electroporation always first occurs in the regions of the membrane where the ITV is the highest.
(iii) Cells in clusters behave differently than isolated cells and their behaviour is dependent on the electric field parameters.
(iv) Under specific circumstances we can observe a domino effect on cells in clusters (cells electroporate one after another, like dominos)
Within the projec we developed three new methods:
(i) A method which allows the construction of realistic numerical models of clusters of cells from a set of microscopic cross-section images. This provides a possibility to determine the ITV on the same cell clusters on which an experiment is carried out, which is of considerable importance in understanding and interpretation of the data.
(ii) A method for replacing cell membranes in the model of a cluster with boundary conditions assigned to the interfaces between the cells and between their interiors and exteriors. This considerably reduces the difficulties associated with construction of models of clusters of cells, and also decreases the number of mesh elements and the time required to solve the problem.
(iii) A method for constructing realistic time-dependent models of cells and cell clusters. The models allow for calculation of the time-dependent changes of the ITV and electroporation. They present an improvement to the existing sequential models of electroporation, because they simulate electroporation in real time.
Significance for the country
The research was performed at the Faculty of Electrical Engineering, University of Ljubljana, which enabled the transfer of knowledge acquired during the course of the project (e.g. the method for construction of realistic numerical models of biological cells, measurements of the induced transmembrane voltage) directly into the educational process. Besides, some of the graduate and postgraduate students actively participated in the research (e.g. visiting postgraduate student Leila Towhidi). Within the International Workshop and Postgraduate School on Electroporation, organized by the Laboratory of Biocybernetics, we demonstrated the method for monitoring the course of electroporation of cell membranes. In the practical example we demonstrated how the method can be used to determine the influence of cell shape and cell orientation on the efficiency of electroporation. The international school enabled a worldwide transfer of knowledge and findings acquired during the project, which increased the reputation of the principal investigator, the group, the University, and the Republic of Slovenia.
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