Basic and Translational SciencePLK-1 Silencing in Bladder Cancer by siRNA Delivered With Exosomes
Section snippets
Cell Culture
UMUC3, SW780, and HEK293 cell lines were purchased from ATCC and cultured in Dulbecco's modified eagle medium containing 10% fetal bovine serum, 100 units/mL penicillin, 100 µg/mL streptomycin, and 2 mmol/L L-glutamine. Normal urothelial cells were purchased from Zen-Bio (Research Triangle Park, NC) and cultured in CnT-Prime Epithelial Culture Medium (Zen-Bio). Cells tested negative for mycoplasma contamination using the MycoAlert Plus Mycoplasma detection kit (Lonza) and they were not cultured
Internalization of HEK293 Exosomes by Bladder Cancer Cells and Normal Urothelial Cells
With the idea of ultimately using exosomes as an intravesical therapy, we investigated the rate of uptake of HEK293 exosomes in bladder cancer cells vs normal urothelial cells. PKH-26-labeled HEK293 exosomes were analyzed on the ImageStreamX after incubating for 6 hours with bladder cells lines UMUC3 and SW780, or with normal urothelial cells. We found that the bladder cancer cells internalize HEK293 exosomes more avidly than normal urothelial cells. Both bladder cancer cell lines tested took
Discussion
Genetic therapeutics have been limited by the availability of an efficient delivery vector. The delivery of siRNA into a cell's cytosol is challenging because it is rapidly degraded, and their size and negative charge limit cellular uptake.13 In addition, siRNA can elicit a powerful immune response, especially with repeated administration. The encapsulation of nucleic acid therapeutics into delivery vectors is a promising strategy to overcome these challenges.13 The ideal delivery vector has
Conclusion
Exosomes have many natural properties that make them an ideal delivery vector for genetic therapeutics. This study adds to the growing body of literature showing the successful use of exosomes as delivery vectors, and it is the first study evaluating their use in bladder cancer with implications in the intravesical therapy setting. Future directions will include alternative gene targets and in vivo models.
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Financial Disclosure: The authors declare that they have no relevant financial interests.
Funding Support: This work was supported in part by the 2013 Urology Care Foundation Residency Research Award to Kristin A. Greco.