Bovine-derived cultured cells, including Madin-Darby bovine kidney cells, are used worldwide; however, lipofection tend to result in low transfection efficiency, which has impeded the progress of veterinary research. We performed experiments to confirm the lipofection efficiency of bovine-derived cultured cells, to identify cells that suitable for lipofection. Several bovine tissues (endometrium, testis, ear tissue and foetal muscle) were collected, and primary cultured cells were prepared. Lipofection assay showed that only bovine endometrium (BE)-derived cells could be transfected efficiently (50‒70%). BE cells can be divided into at least two types of cell populations (BE-1 and BE-2). The BE-1 cells, which were suitable for lipofection, were obtained by passages at short intervals and were negative for cytokeratin- and positive for vimentin-expression; the BE-2 cells did not have these characteristics and were not suitable for lipofection.
Fig: Transfection efficiency of lipofection in bovine-derived cultured cells. (A) MDBK, 293T, BE, BT, BF and BFM cells were seeded on a 24-well culture plate and transfected with 250 ng/well of ZsGreen1-expression vector, using the TransFectin Lipid Reagent. The transfected cells were observed under a fluorescence microscope at 48 h post transfection (hpt; left pictures). The ZsGreen1-positive cell rate and cell viability were measured at 48 hpt (right graph). Asterisks indicate significant differences from the control MDBK cells (***P < 0.001). (B) One BE cell clone could be passaged 30 times and maintained high lipofection efficiency. The transfection assay was described for (A).
Furthermore, the BE-1 cells and artificially immortalised cells of BE-1, iBE-1 cells, were utilised in a reporter assay requiring the introduction of multiple DNAs. Endometrial tissues can be collected from living cows, and BE-1 cells can be obtained easily by controlling passaging timing. The production of BE-1 cells and sharing the methods required to prepare them will contribute to the development of veterinary research.
Shiokawa, M., Miura, R., Okubo, A. et al. Bovine endometrium-derived cultured cells are suitable for lipofection. Sci Rep 11, 16207 (2021). https://doi.org/10.1038/s41598-021-95848-0