Stabilization of gene expression and cell morphology after explant recycling during fin explant culture in goldfish.
The development of fin primary cell cultures for in vitro cellular and physiological studies is hampered by slow cell outgrowth, low proliferation rate, poor viability, and sparse cell characterization. Here, we investigated whether the recycling of fresh explants after a first conventional culture could improve physiological stability and sustainability of the culture. The recycled explants were able to give a supplementary cell culture showing faster outgrowth, cleaner cell layers and higher net cell production. The cells exhibited a highly stabilized profile for marker gene expression including a low cytokeratin 49 (epithelial marker) and a high collagen 1a1 (mesenchymal marker) expression. Added to the cell spindle-shaped morphology, motility behavior, and actin organization, this suggests that the cells bore stable mesenchymal characteristics. This contrast with the time-evolving expression pattern observed in the control fresh explants during the first 2 weeks of culture: a sharp decrease in cytokeratin 49 expression was concomitant with a gradual increase in col1a1. We surmise that such loss of epithelial features for the benefit of mesenchymal ones was triggered by an epithelial to mesenchymal transition (EMT) process or by way of a progressive population replacement process. Overall, our findings provide a comprehensive characterization of this new primary culture model bearing mesenchymal features and whose stability over culture time makes those cells good candidates for cell reprogramming prior to nuclear transfer, in a context of fish genome preservation.
INRA, UR1037 Fish Physiology and Genomics, Campus de Beaulieu, F-35000 Rennes, France.
Nathalie
N
Chenais
INRA, UR1037 Fish Physiology and Genomics, Campus de Beaulieu, F-35000 Rennes, France.
Jean-Jacques
JJ
Lareyre
INRA, UR1037 Fish Physiology and Genomics, Campus de Beaulieu, F-35000 Rennes, France.
Pierre-Yves
PY
Le Bail
INRA, UR1037 Fish Physiology and Genomics, Campus de Beaulieu, F-35000 Rennes, France. Electronic address: catherine.labbe@rennes.inra.fr.
Catherine
C
Labbe
D058500:Animal Fins / Q000166:cytology*; D000818:Animals; D002465:Cell Movement / Q000235:genetics; D049109:Cell Proliferation; D002478:Cells, Cultured; D024042:Collagen Type I / Q000096:biosynthesis; D000090602:Collagen Type I, alpha 1 Chain; D058750:Epithelial-Mesenchymal Transition / Q000502:physiology*; D015870:Gene Expression; D006054:Goldfish / Q000235:genetics*; D018398:Homeodomain Proteins / Q000096:biosynthesis; D007633:Keratins / Q000096:biosynthesis; D050814:Octamer Transcription Factor-3 / Q000096:biosynthesis; D061251:Primary Cell Culture / Q000379:methods*; D016271:Proto-Oncogene Proteins c-myc / Q000096:biosynthesis; D055748:SOXB1 Transcription Factors / Q000096:biosynthesis
D024042:Collagen Type I; D000090602:Collagen Type I, alpha 1 Chain; D018398:Homeodomain Proteins; D050814:Octamer Transcription Factor-3; D016271:Proto-Oncogene Proteins c-myc; D055748:SOXB1 Transcription Factors; D007633:Keratins
Cell culture; Collagen; Cytokeratin; Epithelial; Mesenchymal