Inactivation of the FGF-4 Gene in Embryonic Stem Cells Alters the Growth and/or the Survival of Their Early Differentiated Progeny

• Published in: Developmental biology 1997-12, Vol.192 (2), p.614-629
• Main Authors: Wilder, Phillip J., Kelly, David, Brigman, Kristen, Peterson, Cynthia L., Nowling, Tamara, Gao, Qing-Sheng, McComb, Rodney D., Capecchi, Mario R., Rizzino, Angie
• Format: Article
• Language: English
• Subjects: Animals; Apoptosis; Cell Differentiation - drug effects; Cell Lineage; Cell Survival; Cells, Cultured; Chimera; Female; Fibroblast Growth Factor 4; Fibroblast Growth Factors - genetics; Fibroblast Growth Factors - physiology; Gene Targeting; Heterozygote; Male; Mice; Mice, Inbred C57BL; Proto-Oncogene Proteins - genetics; Proto-Oncogene Proteins - physiology; RNA, Messenger - biosynthesis; Stem Cells - drug effects; Stem Cells - metabolism; Tretinoin - pharmacology
• ISSN: 0012-1606; 1095-564X
• DOI: 10.1006/dbio.1997.8777

Previous studies have shown that early mouse embryos with both FGF-4 alleles inactivated are developmentally arrested shortly after implantation. To understand the roles of FGF-4 during early development, we prepared genetically engineered embryonic stem (ES) cells, which are unable to produce FGF-4. Specifically, we describe the isolation and characterization of ES cells with both FGF-4 alleles inactivated. The FGF-4−/−ES cells do not require FGF-4 to proliferatein vitro,and addition of FGF-4 to the medium has little or no effect on their growth. Thus, FGF-4 does not appear to act as an autocrine growth factor for cultured ES cells. We also demonstrate that FGF-4−/−ES cells, like their unmodified counterparts, are capable of forming highly complex tumors in syngeneic mice composed of a wide range of differentiated cells types, including neural tissue, glandular epithelium, and muscle. In addition, we demonstrate that the FGF-4−/−ES cells can differentiatein vitroafter exposure to retinoic acid; however, the growth and/or survival of the differentiated cells is severely compromised. Importantly, addition of FGF-4 to the culture medium dramatically increases the number of differentiated cells derived from the FGF-4−/−ES cells, in particular cells with many of the properties of parietal extraembryonic endoderm. Finally, we demonstrate that there are differences in the RNA profiles expressed by the differentiated progeny formedin vitrofrom FGF-4−/−ES cells and FGF-4+/+ES cells when they are cultured with FGF-4. Taken together, the studies described in this report indicate that certain lineages formedin vitroare affected by the inactivation of the FGF-4 gene, in particular specific cells that form during the initial stage of ES cell differentiation. Thus, ES cells with both FGF-4 alleles inactivated should shed light on the important roles of FGF-4 during the early stages of mammalian development and help determine why FGF-4−/−embryos die shortly after implantation.