Abstract:SummaryIn vivo electrotransfer is a physical method of gene delivery in various tissues and organs, relying on the injection of a plasmid DNA followed by electric pulse delivery. The importance of the association between cell permeabilization and DNA electrophoresis for electrotransfer efficiency has been highlighted. In vivo electrotransfer is of special interest since it is the most efficient non-viral strategy of gene delivery and also because of its low cost, easiness of realization and safety. The potenti… Show more
“…8 For nonviral transfection, the naked DNA can be delivered into cells or tissues either by chemical carriers such as liposome and cationic polymers or by physical/ mechanical means exemplified by electroporation. 9,10 Effective use of these methods is limited by the low transfection efficiency, the transient expression of gene product and the lack of organ and tissue specificity. Previous studies have suggested that sonoporation (transient increase in cell membrane permeability by ultrasonic exposure) could be an alternative approach as it is not only rapid, reproducible and relatively inexpensive but also provides a safe, noninvasive and efficient means suitable for in vivo gene delivery.…”
Gene transfer into the peritoneal cavity by nonviral methods may provide an effective therapeutic approach for peritoneal diseases. Herein, we investigated the feasibility and the effectiveness of ultrasound-microbubble-mediated delivery of naked plasmid DNA into the peritoneal cavity in rats. Following the intraperitoneal or the intravenous administration of a mixture of plasmid DNA (100 mg) and ultrasound contrast agent microbubbles, an ultrasound transducer was applied on the abdominal wall. The reporter pTRE plasmid encoding Smad7 was used to evaluate transfection efficiency. Smad7 expression was induced by doxycycline in drinking water. We detected less than 10% apoptotic cells and no inflammatory reaction in peritoneal tissues following the ultrasound-microbubble-mediated transfection. More importantly, the insonation significantly improved the transfection efficiency in peritoneal tissues. The transfection efficiency by intraperitoneal delivery route was higher than the intravenous route. The reporter gene, pTRE-Smad7, was readily detected in the parietal peritoneum, mesentery, greater omentum and adipose tissue. The peak of transgene expression occurred 2 days after transfection and the transgene expression diminished in a time-dependent manner thereafter. Overall, the effectiveness and simplicity of the ultrasound-microbubble-mediated system may provide a promising nonviral means for improving gene delivery for treating peritoneal diseases in vivo.
“…8 For nonviral transfection, the naked DNA can be delivered into cells or tissues either by chemical carriers such as liposome and cationic polymers or by physical/ mechanical means exemplified by electroporation. 9,10 Effective use of these methods is limited by the low transfection efficiency, the transient expression of gene product and the lack of organ and tissue specificity. Previous studies have suggested that sonoporation (transient increase in cell membrane permeability by ultrasonic exposure) could be an alternative approach as it is not only rapid, reproducible and relatively inexpensive but also provides a safe, noninvasive and efficient means suitable for in vivo gene delivery.…”
Gene transfer into the peritoneal cavity by nonviral methods may provide an effective therapeutic approach for peritoneal diseases. Herein, we investigated the feasibility and the effectiveness of ultrasound-microbubble-mediated delivery of naked plasmid DNA into the peritoneal cavity in rats. Following the intraperitoneal or the intravenous administration of a mixture of plasmid DNA (100 mg) and ultrasound contrast agent microbubbles, an ultrasound transducer was applied on the abdominal wall. The reporter pTRE plasmid encoding Smad7 was used to evaluate transfection efficiency. Smad7 expression was induced by doxycycline in drinking water. We detected less than 10% apoptotic cells and no inflammatory reaction in peritoneal tissues following the ultrasound-microbubble-mediated transfection. More importantly, the insonation significantly improved the transfection efficiency in peritoneal tissues. The transfection efficiency by intraperitoneal delivery route was higher than the intravenous route. The reporter gene, pTRE-Smad7, was readily detected in the parietal peritoneum, mesentery, greater omentum and adipose tissue. The peak of transgene expression occurred 2 days after transfection and the transgene expression diminished in a time-dependent manner thereafter. Overall, the effectiveness and simplicity of the ultrasound-microbubble-mediated system may provide a promising nonviral means for improving gene delivery for treating peritoneal diseases in vivo.
“…Indeed, a wide range of tissues have been studied including skin, kidney, lung, liver, skeletal and cardiac muscle, joints, spinal cord, brain, retina, cornea and the vasculature. [4][5][6][7] In most studies, electroporation increased gene expression by 100-to 1000-fold compared to injection of naked plasmid DNA. The exact mechanism by which delivery of plasmid into cells is enhanced is not certain, although it is clear that membranes become effectively permeable once a critical voltage has been achieved (in the order of 200 V/cm in vivo).…”
Section: Application Of An Electrical Field Dramatically Enhances Plamentioning
Over the last 5 years, physical methods of plasmid delivery have revolutionized the efficiency of nonviral gene transfer, in some cases reaching the efficiencies of viral vectors. In vivo electroporation dramatically increases transfection efficiency for a variety of tissues. Other methods with clinical precedent, pressure-perfusion and ultrasound, also improve plasmid gene transfer. Alternatives such as focused laser, magnetic fields and ballistic (gene gun) approaches can also enhance delivery. As plasmid DNA appears to be a safe gene vector system, it seems likely that plasmid with physically enhanced delivery will be used increasingly in clinical trials.
“…1 Some applications of ET now envisioned are the endogenous synthesis of EPO, 2 of the soluble receptor to TNFa 3 and of the antiinflammatory cytokine IL-10, 4,5 as well as tumour gene therapy, 6,7 restoration of muscle structural proteins, 8 and DNA vaccination. 9 ET is also applied to tumour electrochemotherapy, 10 where protocols with high voltage and short duration pulses allow antitumoral drugs to be locally delivered to tumour cells.…”
In vivo gene electrotransfer (ET) is a simple method of gene delivery in various tissues relying on the injection of plasmid DNA followed by application of electric pulses. Noninvasive tools are needed to evaluate the ET efficiency and the resulting tissue damages. In this study, we performed ET of rat tibialis muscle after injection of either a plasmid coding for luciferase or a contrast agent (CA) detected by using magnetic resonance imaging (MRI). Plasmid expression and CA intracellular trapped quantity were compared throughout the electric field intensity range 0-300 V/cm. Although the CA trapped quantity reflects only the electropermeabilization step, both measurements were correlated. MRI measurements gave easy access to tridimensional visualization of the labelled zones where the CA has been injected and the applied electric field had a value allowing permeabilization. We also performed MRI measurements of the water transverse relaxation time T2 as an indicator of tissue modification, and tested whether another CA specific for necrosis could be used to detect muscle necrosis at high electric field intensity. In conclusion, MRI measurements may bring multiparametric information upon the efficiency and tissue toxicity of an ET protocol by using a simple and safe CA.
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