A novel method of gene transduction to the murine endometrium using in vivo electroporation

Abstract

To investigate the molecular pathways involved in successful embryo implantation in mammals, we developed a novel method for gene transduction into the murine endometrium using in vivo electroporation. Plasmid DNA with an enhanced green fluorescence protein (EGFP) gene was injected into the uterine cavity of non-pregnant female mice, and electrical pulses were subsequently applied to the uterine horn using plate electrodes. EGFP expression was found only in the uterine luminal epithelium (LE), but not in the stroma. EGFP fluorescence in the LE was limited to the site where the positive side of the electrodes was placed during electric stimulation. These results demonstrated that our novel method enabled us to transduce a gene into a desired location of the murine uterus.

Keywords: EGFP; embryo implantation; in vivo electroporation; uterus.

Fig. 1.

The gene transduction using in vivo electroporation. (A) The line graph of electric pulses by in vivo electroporation. Electric pulses consisted of three poring pulses (PP), three positive transfer pulses (TP+) and three negative transfer pulses (TP-). (B) In vivo electroporation with solely platinum plate electrode. PE; positive electrode, NE; negative electrode, U; uterus. (C) The schema of electroporation. Yellow allow indicates the predictive direction of the electric current. LE; luminal epithelium.

Fig. 2.

EGFP expression in the uterine luminal epithelium (LE). The highly effective transfection of EGFP gene in the LE cells was observed when 3 sets of electric pulses were performed (A), compared with that when only 1 set of pulses (B), or only 1 set of poring pulse (C) was performed. The uterus injected with DNA plasmid without electroporation did not show any EGFP fluorescence (D). PP, poring pulse; TP, transfer pulse; EP, electroporation. Scale bar: 400 µm.

Fig. 3.

One side of the luminal epithelium (LE) shows EGFP fluorescence. (A) The schema of electroporation where the electrical stimuli were applied at three adjacent points along the longitudinal axis of the uterine horns. Yellow allow indicates the predictive direction of the electric current. (B, C) Both samples showed EGFP fluorescence at one side of the LE cells that were placed with positive paddle of electrode. White arrow shows the boundary between the LE and the stroma. Scale bar: 400 µm.