Zhe Wu PhD, Presenter: Employee, General Electric Company

Wenjin Cui, Abstract Co-Author: Nothing to Disclose

To optimize green fluescence protein (GFP) genes delivery to neural progenitor cells with the assistance of ultrasound and microbubbles.

Neural progenitor cells (NPCs) have an inherent tumor tropism, which can be used as a delivery vehicle to target therapeutic gene to tumors. We proposed a new method for transfer gene to mouse NPCs with microbubbles and ultrasound. The NPCs were grown in DMEM supplemented with 20% FBS, 10% horse serum and penicillin (100units/ml)/streptomycin (100μg/ml). Then they were seeded into three OptiCellTM units and grown until 90% confluency. Cationic microbubbles containing DSPC and DSTAP lipids in a 90:10 molar ration were prepared. 20 µg pEGFP-N1 was incubated with microbubbles for 1 hour at room temperature before added into the culture. Opticell plates were turned upside down to allow the microbubbles rising against the cell layer. After 12 hours, the Opticell plates were flipped back. Free bubbles were washed away with PBS. A Sonicconcepts high intensity ultrasound transducer was used to generate the acoustic field to promote the gene delivery in a process known as sonoporation. The ultrasound intensity was monitored by a hydrophone in terms of the peak negative pressure (PNP). Each opticell chamber was evenly divided into 5*5 regions where 5 different ultrasound levels (0KPa, 100KPa, 200KPa, 300KPa, 500KPa) and 5 duty cycles (10% 20% 30% 40% 50%) were applied to these regions respectively. The 0KPa row acted as our control group. Ultrasound frequency was fixed at 1MHz. Each region was slowly translated across the ultrasound focus to receive uniform ultrasound exposure for a total of 60 seconds. This experiment was repeated three times. The transfected cells are further incubated for 36 hours and then observed with phase contrast and florescence microcopy.

No green florescence was observed in the control group where no ultrasound was applied. The maximum transfection efficiency was observed in the 300KPa 40% group. Cell viability was higher than 70% for all groups.

Ultrasound was observed to promote gene transfection to NPCs. Optimized ultrasound parameters were reported.

Due to their inherent tumor tropism, NPCs may be engineered to be therapeutic vehicles once transfected with therapeutic genes. Ultrasound offers potential to transfect cells at designated locations

Wu, Z, Cui, W, Ultrasound-assisted Gene Delivery to Mice Neural Progenitor Cells in the Presence of Microbubbles.  Radiological Society of North America 2011 Scientific Assembly and Annual Meeting, November 26 - December 2, 2011 ,Chicago IL. http://archive.rsna.org/2011/11005826.html