Pressure-Mediated Transfection of Murine Spleen and Liver

Abstract

Extension of in vivo nucleic acid transfection techniques and increased information about those transfection properties and side effects are urgently needed to advance biological research and drug therapy. Tissue pressure-mediated transfection, involving lightly pressing the target tissue after intravenous injection of plasmid DNA or small-interfering RNA (siRNA), is a promising approach because of its high transfection efficiency and resulting low tissue damage. In this study, the gene expression/silencing properties and proinflammatory cytokine production associated with tissue pressure-mediated transfection were evaluated to extend its application. We have found that tissue pressure-mediated transfection can be applied to plasmid DNA and siRNA transfection to the spleen and siRNA transfection to the liver. In addition, we have demonstrated that these methods induce little production of proinflammatory cytokines, including tumor necrosis factor-alpha, interleukin (IL)-6, IL-12, and interferon-gamma. Moreover, we succeeded in controlling and quantifying the degree of pressure on the spleen and kidney and found that 0.59 N/cm(2) is sufficient for efficient and highly reproducible plasmid DNA transfection to the spleen and kidney in mice. Tissue pressure-mediated transfection of the kidney, liver, and spleen exhibits well-balanced characteristics including (1) simple and convenient manipulation, (2) tissue-specific, effective broad transfection properties, and (3) a low inflammatory response. Therefore, this information could be useful for a molecular-level mechanism analysis of diseases at an individual level in mammals, exploration of therapeutic target molecules and evaluation of gene therapy and nucleic acid-based therapy approaches, as well as potential clinical applications.

Liver Suction-Mediated Transfection in Mice Using a Pressure-Controlled Computer System

“…Previous studies demonstrated that the degree of pressure could be used to control the expression levels of the transfected plasmid DNA (pDNA). [9][10][11] Regarding kidneys in mice, renal dysfunction 8) and secretion of proinflammatory cytokines 9) were not observed under the experimental conditions of the tissue pressure method. Recently, Oba et al transfected the miR-200 family of microRNA to mice kidneys via the tissue pressure method and reported that that transfection ameliorated renal tubulointerstitial fibrosis in the mice.…”

“…[8][9][10] In this method, the target organs of the mice are pressed directly after intravenous injection (i.v.) of the naked nucleic acids.…”

Key Physiological Phenomena Governing Transgene Expression Based on Tissue Pressure-Mediated Transfection in Mice

“…10) We have investigated tissue pressure-mediated transfection in order to develop a naked plasmid DNA transfection method with a high gene transfection efficiency and which causes little tissue injury. 11,12) This transfection is performed by light and controlled pressure of the target tissue after normal intravenous injection of plasmid DNA. It is a development that is based on the phenomena described by Liu and Huang in their pioneering report where liver-specific gene expression could be obtained without liver toxicity by massaging the abdomen after intravenous injection of plasmid DNA in mice.…”

“…10) We have been successful in quantitatively controlling the degree of pressure on tissues, and have suggested that plasmid DNA and small-interfering RNA (siRNA) are very efficiently transfected into the murine kidney, liver and spleen without marked tissue damage and this includes an increase in biomarker and pro-inflammatory cytokine production. 11,12) Understanding the key physiological phenomena affecting transgene expression is very important for the proper use of this method and to allow improvements to be made. In general, there are many barriers to gene expression following administration of plasmid DNA: biodistribution, cellular and nuclear translocation of plasmid DNA and transcription and translation.…”

Combination of Nanoparticles with Physical Stimuli toward Cancer Therapy

“…38,39) As more mild physical stimuli, pressure-mediated transfections of intravenously delivered naked pDNA to the kidney, liver and spleen have been reported. 40,41) Moreover, tissue suction using a device enables site-specific transfection by naked pDNA to the kidney, liver, spleen, and heart. 42) On the other hand, rubbing stimuli against the gastric serosal surface greatly enhanced naked pDNA transfer.…”