Mouse fibroblasts are reprogrammed to Oct-4 and Rex-1 gene expression and alkaline phosphatase activity by embryonic stem cell extracts.
A recent remarkable study has shown that when mouse NIH-3T3 fibroblasts are exposed to an embryonic stem cell (ESC) extract, the majority of them expresses the Oct-4 gene, form ESC-like colonies, and embryoid-like bodies that differentiate into cells of the three germ layers. The use of cell extracts for inducing cell dedifferentiation could be a powerful system to obtain large quantities of pluripotent cells. It is thus of crucial importance that the robustness of this method of cell transdifferentiation is tested by other laboratories before it is advanced to a more ambitious use in cell therapy programs. We report here our experimental observations using the same reprogramming protocol on STO and NIH-3T3 mouse fibroblasts. Three are the main results: first, we confirmed an enduring reprogramming activity of the ESC extract, although on a much smaller number of cells that varies from approximately 0.003 to 0.04% of the total population of fibroblasts and with an effect limited to the induction of Oct-4 and Rex-1 gene expression and alkaline phosphatase activity. Second, the expression of OCT-4, SSEA-1, and Forssman antigen proteins was never detected. Third, our work has clearly demonstrated that ESCs may survive the procedure of extract preparation, may be source of contamination that is expanded in culture and give false positive results.
Neri
Tui
T
Laboratorio di Biologia dello Sviluppo, Dipartimento di Biologia Animale, Universita' di Pavia, Italy.
Monti
Manuela
M
Rebuzzini
Paola
P
Merico
Valeria
V
Garagna
Silvia
S
Redi
Carlo Alberto
CA
Zuccotti
Maurizio
M
D000469:Alkaline Phosphatase / Q000378:metabolism*; D000818:Animals; D002454:Cell Differentiation; D002457:Cell Extracts; D002460:Cell Line; D002478:Cells, Cultured; D065150:Cellular Reprogramming; D053595:Embryonic Stem Cells / Q000378:metabolism / Q000502:physiology*; D005347:Fibroblasts / Q000166:cytology* / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D016256:Lewis X Antigen / Q000378:metabolism; D051379:Mice; D050814:Octamer Transcription Factor-3 / Q000096:biosynthesis*; D014157:Transcription Factors / Q000096:biosynthesis*
D002457:Cell Extracts; D016256:Lewis X Antigen; D050814:Octamer Transcription Factor-3; C513672:Zfp42 protein, mouse; D014157:Transcription Factors; D000469:Alkaline Phosphatase