Chapter 8
Nanoelectroporation and Collection of Genetically Modified
Exosomes in Primary Cultures of Dendritic Cells
Silvia Duarte-Sanmiguel, Natalia Higuita-Castro,
and Daniel Gallego-Perez
Abstract
Dendritic cells (DCs) are cells of the immune system that behave as antigen presenters and assist in T cell
activation. DCs have recently been used in cell-based immunotherapies for the treatment of different
diseases due to the lack of adverse nonspecific immune responses, typically elicited by other approaches.
Genetically modified DCs, for example, have been used to stimulate CD4/CD8 antigen presenting
immune responses. However, genetic manipulation of primary DCs remains a challenge. Here we describe
a protocol for nonviral, benign transfection of primary DCs using nanochannel-based electroporation, and
the subsequent collection of genetically modified exosomes for downstream applications.
Key words Dendritic cells, Cell-based immunotherapies, Nanochannel-based electroporation
1 Introduction
Gene therapies are based on the introduction of new genetic material into target cells for the treatment of numerous conditions
[1]. Recent gene therapy studies have focused on different types
of immune cells, including natural killer (NK), lymphocytes, T cells,
and dendritic cells (DCs), depending on the specific cell function
and therapeutic need [2]. DCs, in particular, have attracted a great
deal of attention for this purpose. Their ability to stimulate an
antigen-specific immune response, migrate to lymphoid-rich tissue
and activate CD4+ and CD8+ T-cells after the invasion of foreign
pathogens, and their ability to produce large amounts of potentially
therapeutic exosomes, make them uniquely suited for the next
generation of cell-based therapies [3, 4]. Moreover, clinical trials
using DC-based treatment in cancer reported reduction of immunogenicity after intervention. However, despite the great promise,
progress in DC-based therapies has been partially obstructed by
difficulties in genetically manipulating primary DCs. Viral
Shulin Li et al. (eds.), Electroporation Protocols: Microorganism, Mammalian System, and Nanodevice,
Methods in Molecular Biology, vol. 2050, https://doi.org/10.1007/978-1-4939-9740-4_8,
© Springer Science+Business Media, LLC, part of Springer Nature 2020
79
Nanoelectroporation and Collection of Genetically Modified
Exosomes in Primary Cultures of Dendritic Cells
Silvia Duarte-Sanmiguel, Natalia Higuita-Castro,
and Daniel Gallego-Perez
Abstract
Dendritic cells (DCs) are cells of the immune system that behave as antigen presenters and assist in T cell
activation. DCs have recently been used in cell-based immunotherapies for the treatment of different
diseases due to the lack of adverse nonspecific immune responses, typically elicited by other approaches.
Genetically modified DCs, for example, have been used to stimulate CD4/CD8 antigen presenting
immune responses. However, genetic manipulation of primary DCs remains a challenge. Here we describe
a protocol for nonviral, benign transfection of primary DCs using nanochannel-based electroporation, and
the subsequent collection of genetically modified exosomes for downstream applications.
Key words Dendritic cells, Cell-based immunotherapies, Nanochannel-based electroporation
1 Introduction
Gene therapies are based on the introduction of new genetic material into target cells for the treatment of numerous conditions
[1]. Recent gene therapy studies have focused on different types
of immune cells, including natural killer (NK), lymphocytes, T cells,
and dendritic cells (DCs), depending on the specific cell function
and therapeutic need [2]. DCs, in particular, have attracted a great
deal of attention for this purpose. Their ability to stimulate an
antigen-specific immune response, migrate to lymphoid-rich tissue
and activate CD4+ and CD8+ T-cells after the invasion of foreign
pathogens, and their ability to produce large amounts of potentially
therapeutic exosomes, make them uniquely suited for the next
generation of cell-based therapies [3, 4]. Moreover, clinical trials
using DC-based treatment in cancer reported reduction of immunogenicity after intervention. However, despite the great promise,
progress in DC-based therapies has been partially obstructed by
difficulties in genetically manipulating primary DCs. Viral
Shulin Li et al. (eds.), Electroporation Protocols: Microorganism, Mammalian System, and Nanodevice,
Methods in Molecular Biology, vol. 2050, https://doi.org/10.1007/978-1-4939-9740-4_8,
© Springer Science+Business Media, LLC, part of Springer Nature 2020
79