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DNA delivery to ‘ex vivo’ human liver segments

Gene Therapy volume 19, pages 504–512 (2012)Cite this article

Abstract

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 ⩾10 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: 102–104 eGFP DNA copy per 100 pg of total DNA; transcription index: 105–2 × 106 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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Acknowledgements

We thank Mr Alejo Sempere from the Department of Pathology, University of Valencia, for the immunohistochemistry, the SCSIE from the University of Valencia for the electron microscopy and A Gonzalez and ML Lozano from CIEMAT/CIBERER for qPCR analysis. This work has been partially supported by projects from the Generalidad Valenciana (AP-095/09) and the Ministerio de Ciencia e Innovación (SAF2007–64492 and SAF2011–27002).

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Authors and Affiliations

  1. Departamento de Farmacología, Facultad de Medicina, Universidad de Valencia, Valencia, Spain

    M J Herrero, L Sendra, A I Montilla & S F Aliño

  2. Unidad de Farmacología Clínica, Servicio de Farmacia, Hospital Universitario La Fe, Valencia, Spain

    M J Herrero & S F Aliño

  3. Servicio de Cirugía, Hospital Clínico Universitario, Valencia, Spain

    L Sabater

  4. CIEMAT-CIBERER Hematopoiesis and Gene Therapy Division, Madrid, Spain

    G Guenechea

  5. Instituto de Ciencia de Materiales, Universidad de Valencia, Valencia, Spain

    R Abargues

  6. Servicio de Radiología, Hospital Universitario La Fe, Valencia, Spain

    V Navarro

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  1. M J Herrero
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  2. L Sabater
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  3. G Guenechea
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  8. S F Aliño
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Correspondence to S F Aliño.

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Herrero, M., Sabater, L., Guenechea, G. et al. DNA delivery to ‘ex vivo’ human liver segments. Gene Ther 19, 504–512 (2012). https://doi.org/10.1038/gt.2011.144

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