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In vitro transfection of bone marrow-derived dendritic cells with TATp-liposomes.

Pappalardo JS, Langellotti CA, Di Giacomo S, Olivera V, Quattrocchi V, Zamorano PI, Hartner WC, Levchenko TS, Torchilin VP - Int J Nanomedicine (2014)

Bottom Line: TATp-liposomes (TATp-L) used for DC transfection were prepared using TATp derivatized with a lipid-terminated polymer capable of anchoring in the liposomal membrane.Here, we show that the addition of TATp to DNA-loaded liposomes increased the uptake of DNA in DC.The de novo synthesized gD protein was immunologically stimulating when transfections were performed with TATp-L, as indicated by the secretion of interleukin 6.

View Article: PubMed Central - PubMed

Affiliation: Virology Institute, Center for Research in Veterinary and Agronomic Sciences, National Institute for Agricultural Technology (INTA), Hurlingham, BA, Argentina ; National Council for Scientific and Technical Research (CONICET), Autonomous City of Buenos Aires, Argentina ; Center for Pharmaceutical Biotechnology and Nanomedicine, Northeastern University, Boston, MA, USA.

ABSTRACT
Dendritic cells (DC) are antigen-presenting cells uniquely capable of priming naïve T cells and cross-presenting antigens, and they determine the type of immune response elicited against an antigen. TAT peptide (TATp), is an amphipathic, arginine-rich, cationic peptide that promotes penetration and translocation of various molecules and nanoparticles into cells. TATp-liposomes (TATp-L) used for DC transfection were prepared using TATp derivatized with a lipid-terminated polymer capable of anchoring in the liposomal membrane. Here, we show that the addition of TATp to DNA-loaded liposomes increased the uptake of DNA in DC. DNA-loaded TATp-L increased the in vitro transfection efficiency in DC cultures as evidenced by a higher expression of the enhanced green fluorescent protein and bovine herpes virus type 1 glycoprotein D (gD). The de novo synthesized gD protein was immunologically stimulating when transfections were performed with TATp-L, as indicated by the secretion of interleukin 6.

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Fluorescence microscopy of rhodamine-labeled liposomes in DC.Notes: Liposomes were incubated with DC for 10 minutes in serum-free medium. Cells were washed and incubated additionally in complete medium for 60 or 150 minutes (immersion 100×). TATp-L (A), plain-L (B). Reference marker =10 μm.Abbreviations: DC, dendritic cells; plain-L, plain liposomes; TATp-L, TAT peptide liposomes.
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f3-ijn-9-963: Fluorescence microscopy of rhodamine-labeled liposomes in DC.Notes: Liposomes were incubated with DC for 10 minutes in serum-free medium. Cells were washed and incubated additionally in complete medium for 60 or 150 minutes (immersion 100×). TATp-L (A), plain-L (B). Reference marker =10 μm.Abbreviations: DC, dendritic cells; plain-L, plain liposomes; TATp-L, TAT peptide liposomes.

Mentions: This assay allowed the visualization of liposome uptake by cells with liposomes labeled with trace amounts of rhodamine. The liposomes were incubated with DC for 10 minutes. Cells treated with TATp-L showed a strong adherence to liposomes (Figure 3). Furthermore, the penetration of TATp-L into DC was evident from the accumulation of the liposomes in small, well-defined vesicle-like compartments within these cells. Plain-L-treated DC showed faint background surface staining.


In vitro transfection of bone marrow-derived dendritic cells with TATp-liposomes.

Pappalardo JS, Langellotti CA, Di Giacomo S, Olivera V, Quattrocchi V, Zamorano PI, Hartner WC, Levchenko TS, Torchilin VP - Int J Nanomedicine (2014)

Fluorescence microscopy of rhodamine-labeled liposomes in DC.Notes: Liposomes were incubated with DC for 10 minutes in serum-free medium. Cells were washed and incubated additionally in complete medium for 60 or 150 minutes (immersion 100×). TATp-L (A), plain-L (B). Reference marker =10 μm.Abbreviations: DC, dendritic cells; plain-L, plain liposomes; TATp-L, TAT peptide liposomes.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3928453&req=5

f3-ijn-9-963: Fluorescence microscopy of rhodamine-labeled liposomes in DC.Notes: Liposomes were incubated with DC for 10 minutes in serum-free medium. Cells were washed and incubated additionally in complete medium for 60 or 150 minutes (immersion 100×). TATp-L (A), plain-L (B). Reference marker =10 μm.Abbreviations: DC, dendritic cells; plain-L, plain liposomes; TATp-L, TAT peptide liposomes.
Mentions: This assay allowed the visualization of liposome uptake by cells with liposomes labeled with trace amounts of rhodamine. The liposomes were incubated with DC for 10 minutes. Cells treated with TATp-L showed a strong adherence to liposomes (Figure 3). Furthermore, the penetration of TATp-L into DC was evident from the accumulation of the liposomes in small, well-defined vesicle-like compartments within these cells. Plain-L-treated DC showed faint background surface staining.

Bottom Line: TATp-liposomes (TATp-L) used for DC transfection were prepared using TATp derivatized with a lipid-terminated polymer capable of anchoring in the liposomal membrane.Here, we show that the addition of TATp to DNA-loaded liposomes increased the uptake of DNA in DC.The de novo synthesized gD protein was immunologically stimulating when transfections were performed with TATp-L, as indicated by the secretion of interleukin 6.

View Article: PubMed Central - PubMed

Affiliation: Virology Institute, Center for Research in Veterinary and Agronomic Sciences, National Institute for Agricultural Technology (INTA), Hurlingham, BA, Argentina ; National Council for Scientific and Technical Research (CONICET), Autonomous City of Buenos Aires, Argentina ; Center for Pharmaceutical Biotechnology and Nanomedicine, Northeastern University, Boston, MA, USA.

ABSTRACT
Dendritic cells (DC) are antigen-presenting cells uniquely capable of priming naïve T cells and cross-presenting antigens, and they determine the type of immune response elicited against an antigen. TAT peptide (TATp), is an amphipathic, arginine-rich, cationic peptide that promotes penetration and translocation of various molecules and nanoparticles into cells. TATp-liposomes (TATp-L) used for DC transfection were prepared using TATp derivatized with a lipid-terminated polymer capable of anchoring in the liposomal membrane. Here, we show that the addition of TATp to DNA-loaded liposomes increased the uptake of DNA in DC. DNA-loaded TATp-L increased the in vitro transfection efficiency in DC cultures as evidenced by a higher expression of the enhanced green fluorescent protein and bovine herpes virus type 1 glycoprotein D (gD). The de novo synthesized gD protein was immunologically stimulating when transfections were performed with TATp-L, as indicated by the secretion of interleukin 6.

Show MeSH
Related in: MedlinePlus