Dataset Information

In Vitro Effect of Estradiol and Progesterone on Ovine Amniotic Epithelial Cells.

ABSTRACT: Amniotic epithelial cells (AECs), an emerging source of extrafoetal stem cells, have recently attracted attention for their great regenerative potential. Since AEC amplifications are accompanied by the loss of their native epithelial phenotype and by the progressive reduction of relevant biological properties, the issue to be addressed is the development of effective culture protocols. In this context, recently, it has been demonstrated that progesterone (P4) supplementation during ovine AEC (oAEC) expansion could prevent the undesirable epithelial-mesenchymal transition (EMT). In contrast, there is no information to date on the role of the other pregnancy steroids in culture. With this aim, the present study has been designed to clarify the impact of estradiol (E2), alone or in combination with P4 (12.5??M and 25??M), during oAEC amplification. Steroid supplementations were assessed by testing oAEC proliferation, stemness, EMT, and osteogenic or chondrogenic plasticity. The results indicated that EMT can be prevented exclusively in the presence of high doses of P4, while it occurred rapidly in cells exposed to E2 as denoted by protein (cytokeratin-8 and alpha-SMA) and gene expression (vimentin and snail) profiles. Moreover, steroid exposure was able to influence highly oAEC plasticity. Particularly, P4-treated cells displayed a precommitment towards osteogenic lineage, confirmed by the upregulation of OCN, RUNX2, and the greater deposition of calcium nodules. Conversely, P4 exposure inhibited oAEC chondrogenic differentiation, which was induced in E2-treated cells as confirmed by the upregulation of chondrogenesis-related genes (SOX9, ACAN, and COL2A1) and by the accumulation of Alcian blue-positive extracellular matrix. Simultaneously, E2-treated cells remained unresponsive to osteogenic inductive stimuli. In conclusion, media supplementation with high doses of steroids may be adopted to modulate phenotype and plasticity during oAEC amplification. Relevantly, the osteo or chondro steroid-induced precommitment may open unprecedented cell-based therapies to face the unsolved orthopaedic issues related to osteochondral regeneration.

SUBMITTER: Mauro A

PROVIDER: S-EPMC6458847 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Publications

In Vitro Effect of Estradiol and Progesterone on Ovine Amniotic Epithelial Cells.

Mauro Annunziata A Sanyal Hashimita H Canciello Angelo A Berardinelli Paolo P Russo Valentina V Bernabò Nicola N Valbonetti Luca L Barboni Barbara B

Stem cells international 20190327

Amniotic epithelial cells (AECs), an emerging source of extrafoetal stem cells, have recently attracted attention for their great regenerative potential. Since AEC amplifications are accompanied by the loss of their native epithelial phenotype and by the progressive reduction of relevant biological properties, the issue to be addressed is the development of effective culture protocols. In this context, recently, it has been demonstrated that progesterone (P<sub>4</sub>) supplementation during ov ...[more]

PMID: 31049069

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Project description:Glycogen content in mink uterine glandular and luminal epithelia (GE and LE) is maximal during estrus and is depleted before implantation while embryos are in diapause. Uterine glycogen synthesis in vivo is stimulated by estradiol (E2) while its mobilization is induced by progesterone (P4). Nevertheless, treatment of an immortalized mink uterine epithelial cell line (GMMe) with E2 did not affect glycogen production. Interestingly, insulin alone significantly increased synthesis of the nutrient and glycogen content in response to insulin + E2 was greater than for insulin alone. Our objectives were to determine: 1) If insulin receptor protein (INSR) is expressed by mink uterine GE and LE in vivo and if the amount differs between estrus, diapause and pregnancy; 2) if E2, P4 or insulin regulate insulin receptor gene (Insr) expression by GMMe cells, and 3) if E2 and P4 act independently to regulate glycogen metabolism by GMMe cells and/or if their effects are mediated in part through the actions of insulin. The mean (±S.E.) percent INSR content of uterine epithelia was greatest during diapause (GE: 15.65 ± 0.06, LE:16.56 ± 1.25), much less during pregnancy (GE: 2.53 ± 0.60, LE:2.25 ± 0.32) and barely detectable in estrus (GE: 0.03 ± 0.01, LE:0.02 ± 0.01). Glycogen concentrations in GMMe cells increased 10-fold in response to insulin and 20-fold with insulin + E2 when compared to controls. Expression of Insr was increased 2-fold by insulin and insulin + E2 when compared to controls and there was no difference between the two hormone treatments, indicating that E2 does not increase Insr expression in insulin-treated cells. To simulate E2-priming, cells were treated with Insulin + E2 for 24 h, followed by the same hormones + P4 for the second 24 h (Insulin + E2 ? P4) which resulted in Insr and glycogen levels not different from controls. Similarly, cells treated with Insulin + P4 resulted in glycogen concentrations not different from controls. We conclude that the glycogenic actions of E2 on GMMe cells are due to increased responsiveness of the cells to insulin, but not as a result of up-regulation of the insulin receptor. Glycogen mobilization in response to P4 was the result of decreased glycogenesis and increased glycogenolysis occurring concomitantly with reduced Insr expression. Mink uterine glycogen metabolism appears to be regulated in a reproductive cycle-dependent manner in part as a result of the actions of E2 and P4 on cellular responsiveness to insulin.

Dataset Information

In Vitro Effect of Estradiol and Progesterone on Ovine Amniotic Epithelial Cells.

Publications

In Vitro Effect of Estradiol and Progesterone on Ovine Amniotic Epithelial Cells.

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