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Elucidating the Kinetics of Expression and Immune Cell Infiltration Resulting from Plasmid Gene Delivery Enhanced by Surface Dermal Electroporation.

Mendoza JM, Amante DH, Kichaev G, Knott CL, Kiosses WB, Smith TR, Sardesai NY, Broderick KE - Vaccines (Basel) (2013)

Bottom Line: H&E staining of treated skin sections demonstrated an influx of monocytes and granulocytes at the EP site starting at 4 h and persisting up to day 14 post treatment.Immunological staining revealed a significant migration of lymphocytic cells to the EP site, congregating around cells expressing the delivered antigen.These findings may have implications in the future to design efficient DNA vaccination strategies for the clinic.

View Article: PubMed Central - PubMed

Affiliation: Inovio Pharmaceuticals Inc., 1787 Sentry Parkway West, Building 18, Suite 400, Blue Bell, PA 19422, USA.

ABSTRACT
The skin is an attractive tissue for vaccination in a clinical setting due to the accessibility of the target, the ease of monitoring and most importantly the immune competent nature of the dermal tissue. While skin electroporation offers an exciting and novel future methodology for the delivery of DNA vaccines in the clinic, little is known about the actual mechanism of the approach and the elucidation of the resulting immune responses. To further understand the mechanism of this platform, the expression kinetics and localization of a reporter plasmid delivered via a surface dermal electroporation (SEP) device as well as the effect that this treatment would have on the resident immune cells in that tissue was investigated. Initially a time course (day 0 to day 21) of enhanced gene delivery with electroporation (EP) was performed to observe the localization of green fluorescent protein (GFP) expression and the kinetics of its appearance as well as clearance. Using gross imaging, GFP expression was not detected on the surface of the skin until 8 h post treatment. However, histological analysis by fluorescent microscopy revealed GFP positive cells as early as 1 h after plasmid delivery and electroporation. Peak GFP expression was observed at 24 h and the expression was maintained in skin for up to seven days. Using an antibody specific for a keratinocyte cell surface marker, reporter gene positive keratinocytes in the epidermis were identified. H&E staining of treated skin sections demonstrated an influx of monocytes and granulocytes at the EP site starting at 4 h and persisting up to day 14 post treatment. Immunological staining revealed a significant migration of lymphocytic cells to the EP site, congregating around cells expressing the delivered antigen. In conclusion, this study provides insights into the expression kinetics following EP enhanced DNA delivery targeting the dermal space. These findings may have implications in the future to design efficient DNA vaccination strategies for the clinic.

No MeSH data available.


Related in: MedlinePlus

Rapid and persistent monocyte and granulocyte infiltration is detected at the treatment site following EP-enhanced plasmid delivery. Hematoxylin and eosin (H&E) stained histological analysis of skin sections following ID plasmid administration and EP with SEP in guinea pig skin in a time course. Skin biopsies were removed, cryosectioned, H&E stained and visualized under standard light microscopy (10×). An untreated control (no DNA, no EP) and EP alone post 1 h are also shown. A region of the day 7 time point section is enlarged to indicate skin structures and infiltration of monocyte and granulocyte.
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vaccines-01-00384-f004: Rapid and persistent monocyte and granulocyte infiltration is detected at the treatment site following EP-enhanced plasmid delivery. Hematoxylin and eosin (H&E) stained histological analysis of skin sections following ID plasmid administration and EP with SEP in guinea pig skin in a time course. Skin biopsies were removed, cryosectioned, H&E stained and visualized under standard light microscopy (10×). An untreated control (no DNA, no EP) and EP alone post 1 h are also shown. A region of the day 7 time point section is enlarged to indicate skin structures and infiltration of monocyte and granulocyte.

Mentions: The biopsied sections from Figure 2 were additionally stained with H&E to observe the dynamics of infiltration at the site following EP treatment (Figure 4). Unlike infiltration in the muscle following EP, which is generally a relatively slow process, monocytes and granulocytes were observed migrating to the skin treatment site within 4 h. The inflammatory context persisted for 14 days and appeared resolved at day 21. No evidence of necrosis or localized tissue damage as a result of the EP was observed at any time point. To allow closer analysis of the infiltration and annotation of the structural elements in the skin sections, an enlarged view of the day 7 section is included where the sub-structures and stratum of the skin are noted.


Elucidating the Kinetics of Expression and Immune Cell Infiltration Resulting from Plasmid Gene Delivery Enhanced by Surface Dermal Electroporation.

Mendoza JM, Amante DH, Kichaev G, Knott CL, Kiosses WB, Smith TR, Sardesai NY, Broderick KE - Vaccines (Basel) (2013)

Rapid and persistent monocyte and granulocyte infiltration is detected at the treatment site following EP-enhanced plasmid delivery. Hematoxylin and eosin (H&E) stained histological analysis of skin sections following ID plasmid administration and EP with SEP in guinea pig skin in a time course. Skin biopsies were removed, cryosectioned, H&E stained and visualized under standard light microscopy (10×). An untreated control (no DNA, no EP) and EP alone post 1 h are also shown. A region of the day 7 time point section is enlarged to indicate skin structures and infiltration of monocyte and granulocyte.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4494224&req=5

vaccines-01-00384-f004: Rapid and persistent monocyte and granulocyte infiltration is detected at the treatment site following EP-enhanced plasmid delivery. Hematoxylin and eosin (H&E) stained histological analysis of skin sections following ID plasmid administration and EP with SEP in guinea pig skin in a time course. Skin biopsies were removed, cryosectioned, H&E stained and visualized under standard light microscopy (10×). An untreated control (no DNA, no EP) and EP alone post 1 h are also shown. A region of the day 7 time point section is enlarged to indicate skin structures and infiltration of monocyte and granulocyte.
Mentions: The biopsied sections from Figure 2 were additionally stained with H&E to observe the dynamics of infiltration at the site following EP treatment (Figure 4). Unlike infiltration in the muscle following EP, which is generally a relatively slow process, monocytes and granulocytes were observed migrating to the skin treatment site within 4 h. The inflammatory context persisted for 14 days and appeared resolved at day 21. No evidence of necrosis or localized tissue damage as a result of the EP was observed at any time point. To allow closer analysis of the infiltration and annotation of the structural elements in the skin sections, an enlarged view of the day 7 section is included where the sub-structures and stratum of the skin are noted.

Bottom Line: H&E staining of treated skin sections demonstrated an influx of monocytes and granulocytes at the EP site starting at 4 h and persisting up to day 14 post treatment.Immunological staining revealed a significant migration of lymphocytic cells to the EP site, congregating around cells expressing the delivered antigen.These findings may have implications in the future to design efficient DNA vaccination strategies for the clinic.

View Article: PubMed Central - PubMed

Affiliation: Inovio Pharmaceuticals Inc., 1787 Sentry Parkway West, Building 18, Suite 400, Blue Bell, PA 19422, USA.

ABSTRACT
The skin is an attractive tissue for vaccination in a clinical setting due to the accessibility of the target, the ease of monitoring and most importantly the immune competent nature of the dermal tissue. While skin electroporation offers an exciting and novel future methodology for the delivery of DNA vaccines in the clinic, little is known about the actual mechanism of the approach and the elucidation of the resulting immune responses. To further understand the mechanism of this platform, the expression kinetics and localization of a reporter plasmid delivered via a surface dermal electroporation (SEP) device as well as the effect that this treatment would have on the resident immune cells in that tissue was investigated. Initially a time course (day 0 to day 21) of enhanced gene delivery with electroporation (EP) was performed to observe the localization of green fluorescent protein (GFP) expression and the kinetics of its appearance as well as clearance. Using gross imaging, GFP expression was not detected on the surface of the skin until 8 h post treatment. However, histological analysis by fluorescent microscopy revealed GFP positive cells as early as 1 h after plasmid delivery and electroporation. Peak GFP expression was observed at 24 h and the expression was maintained in skin for up to seven days. Using an antibody specific for a keratinocyte cell surface marker, reporter gene positive keratinocytes in the epidermis were identified. H&E staining of treated skin sections demonstrated an influx of monocytes and granulocytes at the EP site starting at 4 h and persisting up to day 14 post treatment. Immunological staining revealed a significant migration of lymphocytic cells to the EP site, congregating around cells expressing the delivered antigen. In conclusion, this study provides insights into the expression kinetics following EP enhanced DNA delivery targeting the dermal space. These findings may have implications in the future to design efficient DNA vaccination strategies for the clinic.

No MeSH data available.


Related in: MedlinePlus