Calcium phosphate-mediated gene delivery using simulated body fluid (SBF).
The present study aimed at developing a new approach in gene delivery of calcium phosphate nanoparticles through simulated body fluid (CaP-SBF). The physicochemical and biological characteristics of the CaP-SBF nanoparticles were compared with those made in pure water (CaP-water) via a similar procedure. The CaP-SBF and CaP-water solutions were then adjusted to two different pH values of 7.4 and 8.0, mixed with plasmid DNA (pDNA), and added in varying amounts to human embryonic kidney (HEK 293T) cells. The transfection efficiency and cell viability were studied in vitro by reporter gene (luciferase and Enhanced Green Fluorescent Protein) expression and the resazurin reduction assay, respectively, 24 and 48 h after the incubation with the nanoparticles. Our results indicated considerably high in vitro transfection efficiency for CaP-SBF/DNA complexes at physiological pH (7.4) with high amounts of CaP. Additionally, the SBF solution exhibited the ability to reduce the rapid growth of CaP particles over time, leading to higher transfection efficiency of CaP-SBF/DNA complexes than those made in water (CaP-water/DNA).
CQM - Centro de Química da Madeira, MMRG, Universidade da Madeira, Campus Universitário da Penteada, 9000-390 Funchal, Portugal. nouri@uma.pt
Alireza
A
Nouri
Rita
R
Castro
José L
JL
Santos
César
C
Fernandes
João
J
Rodrigues
Helena
H
Tomás
D001826:Body Fluids / Q000737:chemistry; D002130:Calcium Phosphates / Q000737:chemistry*; D002470:Cell Survival; D004247:DNA / Q000008:administration & dosage*; D005786:Gene Expression Regulation; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D015316:Genetic Therapy / Q000379:methods; D049452:Green Fluorescent Proteins / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008156:Luciferases / Q000235:genetics; D053758:Nanoparticles; D010957:Plasmids; D013997:Time Factors; D014162:Transfection
D002130:Calcium Phosphates; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D004247:DNA; C020243:calcium phosphate; D008156:Luciferases