Advisor(s)

Mansoor M. Amiji

Contributor(s)

Samuel Gatley, Rebecca L. Carrier

Date of Award

2010

Date Accepted

12-2010

Degree Grantor

Northeastern University

Degree Level

M.S.

Degree Name

Master of Science

Department or Academic Unit

Bouve College of Health Sciences. Department of Pharmaceutical Sciences

Keywords

IL-10, inflammation

Subject Categories

Immunology, Genetics, Emulsions (Pharmacy), Gene therapy, Macrophages, Nanoparticles, Transfection

Disciplines

Pharmacy and Pharmaceutical Sciences

Abstract

Purpose:Multi-compartmental delivery systems can effectively deliver therapeutic payloads by overcoming different extra- and intra-cellular barriers. The main objective of this study was to develop DNA-encapsulated solid nanoparticles-in-emulsion formulation to enhance gene delivery and transfection in macrophages.

Methods:Type B gelatin based nanoparticles were prepared by ethanol water solvent displacement method. Safflower oil-containing water in oil in water multiple emulsion was prepared by a two step emulsification technique with the help of Silverson homogenizer L4RT®. EGFP-N1 plasmid DNA that expresses enhanced green fluorescent protein (GFP) was encapsulated in solid type B gelatin nanoparticles (GNP) which were further encapsulated in the innermost aqueous phase of safflower oil containing water in oil in water (W/O/W) multiple emulsion (ME). Fluorescence microscopy and GFP-specific ELISA were performed for qualitative and quantitative determination of plasmid EGFP-N1 gene transfection in murine adherent alveolar macrophages J774A.1.

Results:Particle size of the NiE formulations was found to be less than or equal to 5 um in diameter. Results obtained from gene transfection evaluations showed that plasmid EGFP-N1 or mIL-10 loaded NiE formulations offered higher and sustained GFP expression compared GFP expression observed with plasmid EGFP-N1 or mLI-10 loaded ME and GNP formulations or plasmid DNA complexed with Lipofectin® , a cationic lipid based standard gene transfection reagent in J774A.1 murine adherent alveolar macrophage cell lines.

Conclusion:The results of this study show that DNA-containing solid gelatin nanoparticles can be encapsulated in the innermost aqueous phase of the W/O/W multiple emulsions to form NiE formulations. EGFP-N1 or mIL-10 plasmid-loaded NiE formulations were capable producing sustained gene transfection in J774A.1 murine adherent alveolar macrophage cell lines. In addition mIL10 transfection using NiE formulation showed highest amount of TNFα and IL-1β suppression upon LPS stimulation.

Document Type

Master's Thesis

Rights Information

copyright 2010

Rights Holder

Husain Attarwala

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