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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

Gene delivery enhanced by dermal electroporation induces sustained expression on the surface of the skin. Time course (1 h post treatment to day 21) of green fluorescent protein (GFP) expression after intradermal (ID) plasmid administration followed by surface electroporation (SEP) in guinea pig skin visualized under natural (top panel) and fluorescent (lower panel) light. An untreated control is also shown. Photos are representative examples of multiple treatments.
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vaccines-01-00384-f001: Gene delivery enhanced by dermal electroporation induces sustained expression on the surface of the skin. Time course (1 h post treatment to day 21) of green fluorescent protein (GFP) expression after intradermal (ID) plasmid administration followed by surface electroporation (SEP) in guinea pig skin visualized under natural (top panel) and fluorescent (lower panel) light. An untreated control is also shown. Photos are representative examples of multiple treatments.

Mentions: To assess the expression kinetics resulting from gene delivery with a dermal EP device, a plasmid expressing GFP was injected into guinea pig skin at defined time points. Surface EP (SEP) was immediately performed after each injection. The animals were sacrificed post treatments and the skin excised and visualized under a fluorescent microscope (Figure 1). GFP expression appeared at 6 h on the surface of the skin and persisted for seven days. The peak expression was observed at 24 h. During the peak times (24–72 h), the expression was robust and matched in size the diameter of the injection bubble (approximately 5 mm). Within the gross localization pattern of GFP, smaller islands of expression were also noted which coincided in shape and spacing with the direct contact the electrodes make with the skin. The biopsies shown in the figure are representative examples of those seen in multiple treatments on multiple animals.


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)

Gene delivery enhanced by dermal electroporation induces sustained expression on the surface of the skin. Time course (1 h post treatment to day 21) of green fluorescent protein (GFP) expression after intradermal (ID) plasmid administration followed by surface electroporation (SEP) in guinea pig skin visualized under natural (top panel) and fluorescent (lower panel) light. An untreated control is also shown. Photos are representative examples of multiple treatments.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

vaccines-01-00384-f001: Gene delivery enhanced by dermal electroporation induces sustained expression on the surface of the skin. Time course (1 h post treatment to day 21) of green fluorescent protein (GFP) expression after intradermal (ID) plasmid administration followed by surface electroporation (SEP) in guinea pig skin visualized under natural (top panel) and fluorescent (lower panel) light. An untreated control is also shown. Photos are representative examples of multiple treatments.
Mentions: To assess the expression kinetics resulting from gene delivery with a dermal EP device, a plasmid expressing GFP was injected into guinea pig skin at defined time points. Surface EP (SEP) was immediately performed after each injection. The animals were sacrificed post treatments and the skin excised and visualized under a fluorescent microscope (Figure 1). GFP expression appeared at 6 h on the surface of the skin and persisted for seven days. The peak expression was observed at 24 h. During the peak times (24–72 h), the expression was robust and matched in size the diameter of the injection bubble (approximately 5 mm). Within the gross localization pattern of GFP, smaller islands of expression were also noted which coincided in shape and spacing with the direct contact the electrodes make with the skin. The biopsies shown in the figure are representative examples of those seen in multiple treatments on multiple animals.

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