Mechanism of plasmid delivery by hydrodynamic tail vein injection. I. Hepatocyte uptake of various molecules

Sebestyén, Magdolna G.; Budker, Vladimir G.; Budker, Tatiana; Субботин, Владимир; Zhang, Guofeng; Monahan, Sean; Lewis, David L.; Wong, So C.; Hagstrom, James E.; Wolff, Jon A.

doi:10.1002/jgm.921

Cited by 92 publications

(82 citation statements)

References 42 publications

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“…The exact mechanism of gene delivery mediated by hydrodynamic injection remains unknown, but two routes of entry of DNA plasmids into hepatocytes have been suggested: plasma membrane rupture 19 and increased membrane permeation, 5,8,20,21 possibly through endocytic vesicles. In the present study we confirm that hydrodynamic injection mediates an increase in sinusoid diameter, with an endothelium that combines both continuous and discontinuous zones, although in no case was generalized disruption observed.…”

Section: Discussionmentioning

confidence: 99%

DNA delivery to ‘ex vivo’ human liver segments

et al. 2011

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Hydrodynamic injection is an efficient procedure for liver gene therapy in rodents but with limited efficacy in large animals, using an 'in vivo' adapted regional hydrodynamic gene delivery system. We study the ability of this procedure to mediate gene delivery in human liver segments obtained by surgical resection. Watertight liver segments were retrogradely injected from hepatic vein with a saline solution containing a plasmid bearing the enhanced green fluorescent protein (eGFP) gene, under different conditions of flow rate (1, 10 and 20 ml s -1 ) and final perfused volume. Samples were cultured for 1 to 2 days and used for microscopy and molecular analysis of gene expression. The fluorescent and immunohistochemistry studies indicated that in segments injected at X10 ml s -1 , good and wide gene expression was present in the liver sections and the molecular analysis reinforced the histological observation in a quantitative manner (index of apparent gene delivery: 10 2 -10 4 eGFP DNA copy per 100 pg of total DNA; transcription index: 10 5 -2Â10 6 eGFP RNA copy per 100 ng of total RNA). In addition, injected gold nanoparticles (15 nm diameter) suggested that DNA delivery to hepatocytes must involve a facilitated permeation process without membrane disruption. In summary, we show that retrograde venous injection of watertight human liver segment is an anadromous procedure that results in wide liver gene delivery and good gene expression. However, additional studies will be necessary to clarify the influence of the prolonged ischemia injury to hepatocytes in our model.

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Section: Discussionmentioning

confidence: 99%

DNA delivery to ‘ex vivo’ human liver segments

et al. 2011

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“…Equal amounts (10 mg) of the pGL3-CMV-Luc reporter plasmid were delivered by i.d. injection into mouse footpad skin (a single injection of the indicated volume was performed at the same pad location on each paw) in different volumes (10,25,50,75 and 100 ml) of PBS vehicle (pressure-fection), and luciferase expression levels were assessed by BLI (Figure 1). This plasmid uses the immediate-early promoter from cytomegalovirus (CMV) as a strong constitutive promoter.…”

Section: Increased Pressure During Hydrodynamic Id Injection Of Plamentioning

confidence: 99%

“…The need for pressure in the skin, however, is in good agreement with hydrodynamic nucleic acid delivery to other organs including liver, kidney and muscle, 15 which may involve perturbation of cellular membranes, such as has been described for hepatocyte uptake following intravascular injection of nucleic acids. 49,50 Several studies have argued against membrane disruption (at least in hepatocytes), and point toward a receptor-mediated mechanism of uptake, where pressure may enhance gene transfer by increasing nucleic acid interactions with cells. 29,36 On the other hand, the combination of electroporation with i.d.…”

Section: Nuclei Are Visualized With Dapi (Blue)mentioning

confidence: 99%

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Increased interstitial pressure improves nucleic acid delivery to skin enabling a comparative analysis of constitutive promoters

González-González

Spitler

et al. 2010

Gene Ther

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Nucleic acid-based therapies hold great promise for treatment of skin disorders if delivery challenges can be overcome. To investigate one mechanism of nucleic acid delivery to keratinocytes, a fixed mass of expression plasmid was intradermally injected into mouse footpads in different volumes, and reporter expression was monitored by intravital imaging or skin sectioning. Reporter gene expression increased with higher delivery volumes, suggesting that pressure drives nucleic acid uptake into cells after intradermal injections similar to previously published studies for muscle and liver. For spatiotemporal analysis of reporter gene expression, a dual-axis confocal (DAC) fluorescence microscope was used for intravital imaging following intradermal injections. Individual keratinocytes expressing hMGFP were readily visualized in vivo and initially appeared to preferentially express in the stratum granulosum and subsequently migrate to the stratum corneum over time. Fluorescence microscopy of frozen skin sections confirmed the patterns observed by intravital imaging. Intravital imaging with the DAC microscope is a noninvasive method for probing spatiotemporal control of gene expression and should facilitate development and testing of new nucleic acid delivery technologies.

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“…Yoshino and associates 20 have developed a surgical protocol based on Eastman's research. Upon the insertion of an occlusion balloon catheter, through the right external jugular vein, into the left hepatic vein under fluoroscopic guidance, the hepatic artery and the portal vein were identified and clamped after a midline incision.…”

Section: Swinementioning

confidence: 99%

Animal Models for Hydrodynamic Gene Delivery

Katsimpoulas¹,

Zacharoulis²,

Habib³

et al. 2012

Chemical Biology

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236The vehicle for the molecules is Normal Saline, Ringer's Solution or Phosphate Buffered Saline and the dosage range from 0.1 to 10 mg/kg, depending on the application. The molecules can be DNA, as plasmid, fragment or artificial chromosome, RNA, single or double stranded, genomic or not, polymers, proteins and small compounds. The main application of the hydrodynamic delivery is the therapy studies, especially genes encoding secretory proteins which can be even isolated and purified. A different area of interest is the gene function analysis in a whole animal instead of a cell culture; the transfection is applied with the hydrodynamic technique and the conclusions are withdrawn from the whole animal. The study of DNA sequences can be achieved with HGD, where promoters, introns, enhancers, suppressors can be delivered intracellular in whole animal. Moreover new animal models are possible to be created for study human diseases, mainly viral due to the fact that they are species specific 5,8,10 .

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Mechanism of plasmid delivery by hydrodynamic tail vein injection. I. Hepatocyte uptake of various molecules

Cited by 92 publications

References 42 publications

DNA delivery to ‘ex vivo’ human liver segments

DNA delivery to ‘ex vivo’ human liver segments

Increased interstitial pressure improves nucleic acid delivery to skin enabling a comparative analysis of constitutive promoters

Animal Models for Hydrodynamic Gene Delivery

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