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Characterization and visualization of the liposecretion process taking place during ceiling culture of human mature adipocytes
Author(s) -
Côté Julie Anne,
Gauthier MarieFrédérique,
Ostinelli Giada,
Brochu Dannick,
Bellmann Kerstin,
Marette André,
Julien François,
Lebel Stéfane,
Tchernof André
Publication year - 2019
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.27931
Subject(s) - biology , microbiology and biotechnology , adipose tissue , mitosis , adipocyte , population , lipid droplet , adipogenesis , cell culture , cell , 3t3 l1 , mesenchymal stem cell , biochemistry , genetics , demography , sociology
Objective To investigate and further characterize the process of mature adipocyte dedifferentiation. Our hypothesis was that dedifferentiation does not involve mitosis but rather a phenomenon of liposecretion. Methods Mature adipocytes were isolated by collagenase digestion of human adipose tissue samples. Ceiling cultures were established using our six‐well plate model. Cells were treated with cytosine β‐ d ‐arabinofuranoside (AraC) or vincristine (VCR), two agents blocking cell division, and were compared with vehicle. Liposecretion events were visualized by time‐lapse microscopy, with and without AraC in adipocytes transducted with a baculovirus. Microscopic analyses were performed after labeling phosphorylated histone 3 and cyclin B1 in ceiling cultures. Results Treatment with AraC almost entirely prevented the formation of fibroblasts up to 12 days of ceiling culture. Similar results were obtained with VCR. The antimitotic effectiveness of the treatment was confirmed in fibroblast cultures from the adipose tissue stromal‐vascular fraction by proliferation assays and colony‐forming unit experiments. Using time‐lapse microscopy, we visualized liposecretion events in which a large lipid droplet was rapidly secreted from isolated mature adipocytes. The same phenomenon was observed with AraC. This was observed in conjunction with histone 3 phosphorylation and cyclin B1 segregation to the nucleus. Conclusion Our results support the notion that dedifferentiation involves rapid secretion of the lipid droplet by the adipocytes with concomitant generation of fibroblast‐like cells that subsequently proliferate to generate the dedifferentiated adipocyte population during ceiling culture. The presence of mitotic markers suggests that this process involves cell cycle progression, although cell division does not occur.