Murine Embryonic Fibroblast Cell Lines Differentiate into Three Mesenchymal Lineages to Different Extents: New Models to Investigate Differentiation Processes

Abstract

Various diseases, injuries, and congenital abnormalities may result in degeneration and loss of organs and tissues. Recently, tissue engineering has offered new treatment options for these common, severe, and costly problems in human health care. Its application is often based on the usage of differentiated stem cells. However, despite intensive research and growing knowledge, many questions remain unresolved in the process of cell differentiation. The aim of this study was to find standardized cell models for analyzing molecular mechanisms of cell differentiation. We investigated the multipotency of three standardized murine embryonic fibroblast cell cultures using histological staining, western blotting, and quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). Our results demonstrated that NIH-3T3 and mouse embryonic fibroblast (MEF) cells were able to differentiate into adipogenic, chondrogenic, and osteogenic lineages expressing typical differentiation markers. Interestingly, Flp-In-3T3 cells did not differentiate into any of the three mesenchymal lineages, although this cell line is genetically closely related to NIH-3T3. The results were confirmed by histological staining. Flp-In-3T3, NIH-3T3, and MEF cells have usually been used for DNA transfections, recombinant protein expression, and as ''feeder cells.'' Unlike mesenchymal stem cells (MSCs) and mesenchymal progenitor cells (MPCs), they are easy to obtain and to expand and are less prone to change their structure and morphology, even at higher passages. Our results suggest that Flp-In-3T3, MEF, and NIH-3T3 cells are highly suitable to be used as models to analyze molecular mechanisms of cell differentiation.

Differentiated mitochondrial function in mouse 3T3 fibroblasts and human epithelial or endothelial cells in response to chemical exposure

“…Fibroblasts are dispersed throughout the connective tissue of the body, and their morphology and physiology have been found to vary depending on the body part . In addition, it is not known which tissues give rise to 3T3 fibroblasts because studies have tended to use homogenized embryo tissue before the preparation of cell culture . Therefore, the comparison of the sensitivity of 3T3 fibroblasts with those of other cell lines represented by the present study offers a valuable contribution to existing knowledge of fibroblast metabolism.…”

Direct conversion of adult human skin fibroblasts into functional Schwann cells that achieve robust recovery of the severed peripheral nerve in rats

“…Also, the use of the umbilical cord as a source of MSCs gives concerns such as limited supply. Although there are some studies that have shown the multi‐potentiality of fibroblasts (Dastagir et al, ; Sudo et al, ), these cells are generally considered to be differentiated, so that, unlike MSCs, their differentiation potential is not considered sufficient for clinical application (Brohem et al, ). In contrast, fibroblasts can be harvested from the skin much more easily than MSCs, and, similar to MSCs, they can be collected from the patient without ethical concerns.…”

Mouse Embryonic Fibroblast Adipogenic Potential: A Comprehensive Transcriptome Analysis

“…Most knowledge of AT is based on in vitro studies of committed murine cell lines, notably 3T3-L1 and 3T3-F442A [28,29], and human cell lines, particularly mesenchymal stem cells (MSCs) [30,31]. However, some studies have demonstrated the adipogenic potential of mouse embryonic fibroblast (MEF) [16,[32][33][34]. MEFs, isolated from 14-day-old mouse embryos (C57/BL6 52 strain), are classically used as 'feeder layers' for human and mouse embryonic stem cells, for DNA transfection assays, recombinant protein expression, and utilization for epigenome and transcriptome analysis (i.e.…”