Establishment and characterization of immortalized mouse embryonic fibroblasts by SV40 large T antigen
Year: 2026
Type: article
Abstract: Mouse embryonic fibroblasts (MEFs), which are typically derived from mouse embryos between embryonic days 12.5 and 13.5, serve as a widely used feeder layer for pluripotent stem cell culture. MEFs exhibit a limited proliferative capacity and are typically used within five passages after which they undergo early senescence. However, the repeated preparation of primary MEFs introduces batch-to-batch variability and raises ethical concerns regarding animal use, thereby limiting the reproducibility and scalability of stem cell culture. To overcome these limitations, here we established a nontumorigenic immortalized MEF line (designated as SV40-CMEF) from C57BL/6J mice via SV40 large T antigen (SV40LT) transfection. SV40-CMEFs showed genomic integration, enhanced proliferation, and the capacity to support the self-renewal of R1/E mouse embryonic stem cells (mESC) as feeder cells. They also retained their tri-lineage differentiation potential. Notably, SV40-CMEFs did not form colonies in soft agar or tumors in immunodeficient mice. Transcriptomic profiling revealed substantial global remodeling compared with that in primary MEFs; however, functional gene modules associated with feeder capacity, fibroblast identity, and proliferation were selectively retained, whereas senescence markers were suppressed. This cell line provides a convenient, well-characterized resource for stem cell culture and cell differentiation research.
Authors: Xin Li, Ting Shao, Lan Xu, Li Zheng, Yue-Lei Chen
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $3,801
Award: G3253879448
OpenAlex ID: W7223183728