Objective To investigate the feasibility of gene transfection into retinal pigment epithelial (RPE) cells and photoreceptors (PRs) in vivo electroporation. Methods A total of 147 Sprague-Dawley (SD) rats were divided into 5, 10, 15, 20, 25, 30 and 35 V group according to different voltage. The right eyes of rats underwent the injection of eukaryotic expressive plasmid of enhanced green fluorescent protein (EGFP) pEGFP-N1 into subretinal space as experimental eyes; the left eyes were injected with TE buffer as control eyes. Each group was divided into RPE and RP subgroups according to different transfection direction. There were same parameters of 99 ms pulse width, 0.5 s pulse interval and 5 consecutive pulses except different voltage in groups. With a negative charge in the electric field was transfected into RPE cell layer, reverse electrode set to be transfected into PR cell layer. Retina mounts were made on seven days after transfection and the fluorescence of EGFP was photographed by fluorescent microscope. The expression of EGFP mRNA and protein were detected by reverse transcription polymerase chain reaction technique (RT-PCR) and Western blot.Results On seven days after transfection, in RPE subgroups, there were no specific fluorescence expressions in RPE cell layer and retina mounts of control eyes, while there were fluorescence expressions in experimental eyes. Western blot showed that the grayscale ratio of EGFP protein and beta;actin protein bands rose with the increased voltage. RT-PCR showed that each group produced positive amplification bands, and the relative ratio of gray level of EGFP mRNA and GADPH mRNA amplified bands gradually increased with the increased voltage.Conclusion Electroporation is an effective method for gene delivery into RPE cells in vivo.
To investigate the effect of membrane thickness on the coupled electroporation–electrodeformation response of the cell membrane, a coupled model incorporating a dynamic Young’s modulus was developed to simulate transmembrane voltage, spatiotemporal pore evolution, and cell deformation at different membrane thicknesses. As membrane thickness increased from 6 to 10 nm, both the transmembrane voltage and pore radius peaked earlier. At 50 ns, pore density at the cell poles increased by 19.78%–67.44%, total electroporated area by 257.48%–859.36%, and the reduction in Young’s modulus became more pronounced. Despite increased membrane softening, peak Maxwell stress decreased by 26.15%–63.28% and remained the dominant factor, reducing peak displacement at the cell poles by 12.35%–34.75%. These findings show that membrane thickness modulates the coupled response by jointly affecting electrically induced pore formation, membrane mechanical properties, and Maxwell stress. The model provides a theoretical basis for designing pulse parameters for cells with different membrane structural characteristics.