Abstract
The development of a multicellular spheroid comprising bovine endometrial epithelial cells (BEE) and bovine endometrial stromal cells (BES) is described in this study. The BES were cultured to confluence in medium with L-ascorbic acid phosphate magnesium salt n-hydrate (AsA-P) which stimulates collagen synthesis in BES. The BEE were co-cultured on a BES cell-sheet for 24 h before detachment of the cell-sheet to generate a hetero-spheroid. After EDTA treatment and agitating with pipette, the floating cell-sheet shrank and became an aggregated cell mass in a few days; it finally formed a round-shaped hetero-spheroid composed of BES and BEE. Histological examination found that hetero-spheroids were covered with BEE on the outer laver. When cell viability was examined with TUNEL (terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling), no positive signal was detected in the spheroid for up to 10 days. Immunofluorescence observations showed that spheroids contained abundant extracellular matrices, including type-I, -III, -IV collagen, fibronectin, and laminin. PGF2α produced by hetero-spheroids in response to oxytocin was significantly higher than those produced by monolayer cultured BEE (P<0.05). MMPs were not detected in media from spheroids cultured for 5 days after detachment of the cell sheet. These results indicate that bovine endometrial cells have the capacity to regenerate as a multicellular spheroid after treatment with ascorbate in vitro. The spheroid displays an endometrium-mimic feature. Thus, we conclude that spheroids formed by BES and BEE are a useful in vitro model of bovine endometrium.
| Original language | English |
|---|---|
| Pages (from-to) | 258-269 |
| Number of pages | 12 |
| Journal | Placenta |
| Volume | 24 |
| Issue number | 2-3 |
| DOIs | |
| Publication status | Published - Jan 1 2003 |
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All Science Journal Classification (ASJC) codes
- Reproductive Medicine
- Obstetrics and Gynaecology
- Developmental Biology
Cite this
A three-dimensional cell culture model for bovine endometrium : Regeneration of a multicellular spheroid using ascorbate. / Yamauchi, N.; Yamada, O.; Takahashi, T.; Imai, K.; Sato, T.; Ito, T.; Hashizume, K.
In: Placenta, Vol. 24, No. 2-3, 01.01.2003, p. 258-269.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - A three-dimensional cell culture model for bovine endometrium
T2 - Regeneration of a multicellular spheroid using ascorbate
AU - Yamauchi, N.
AU - Yamada, O.
AU - Takahashi, T.
AU - Imai, K.
AU - Sato, T.
AU - Ito, T.
AU - Hashizume, K.
PY - 2003/1/1
Y1 - 2003/1/1
N2 - The development of a multicellular spheroid comprising bovine endometrial epithelial cells (BEE) and bovine endometrial stromal cells (BES) is described in this study. The BES were cultured to confluence in medium with L-ascorbic acid phosphate magnesium salt n-hydrate (AsA-P) which stimulates collagen synthesis in BES. The BEE were co-cultured on a BES cell-sheet for 24 h before detachment of the cell-sheet to generate a hetero-spheroid. After EDTA treatment and agitating with pipette, the floating cell-sheet shrank and became an aggregated cell mass in a few days; it finally formed a round-shaped hetero-spheroid composed of BES and BEE. Histological examination found that hetero-spheroids were covered with BEE on the outer laver. When cell viability was examined with TUNEL (terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling), no positive signal was detected in the spheroid for up to 10 days. Immunofluorescence observations showed that spheroids contained abundant extracellular matrices, including type-I, -III, -IV collagen, fibronectin, and laminin. PGF2α produced by hetero-spheroids in response to oxytocin was significantly higher than those produced by monolayer cultured BEE (P<0.05). MMPs were not detected in media from spheroids cultured for 5 days after detachment of the cell sheet. These results indicate that bovine endometrial cells have the capacity to regenerate as a multicellular spheroid after treatment with ascorbate in vitro. The spheroid displays an endometrium-mimic feature. Thus, we conclude that spheroids formed by BES and BEE are a useful in vitro model of bovine endometrium.
AB - The development of a multicellular spheroid comprising bovine endometrial epithelial cells (BEE) and bovine endometrial stromal cells (BES) is described in this study. The BES were cultured to confluence in medium with L-ascorbic acid phosphate magnesium salt n-hydrate (AsA-P) which stimulates collagen synthesis in BES. The BEE were co-cultured on a BES cell-sheet for 24 h before detachment of the cell-sheet to generate a hetero-spheroid. After EDTA treatment and agitating with pipette, the floating cell-sheet shrank and became an aggregated cell mass in a few days; it finally formed a round-shaped hetero-spheroid composed of BES and BEE. Histological examination found that hetero-spheroids were covered with BEE on the outer laver. When cell viability was examined with TUNEL (terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling), no positive signal was detected in the spheroid for up to 10 days. Immunofluorescence observations showed that spheroids contained abundant extracellular matrices, including type-I, -III, -IV collagen, fibronectin, and laminin. PGF2α produced by hetero-spheroids in response to oxytocin was significantly higher than those produced by monolayer cultured BEE (P<0.05). MMPs were not detected in media from spheroids cultured for 5 days after detachment of the cell sheet. These results indicate that bovine endometrial cells have the capacity to regenerate as a multicellular spheroid after treatment with ascorbate in vitro. The spheroid displays an endometrium-mimic feature. Thus, we conclude that spheroids formed by BES and BEE are a useful in vitro model of bovine endometrium.
UR - http://www.scopus.com/inward/record.url?scp=0012900968&partnerID=8YFLogxK
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U2 - 10.1053/plac.2002.0901
DO - 10.1053/plac.2002.0901
M3 - Article
C2 - 12566253
AN - SCOPUS:0012900968
VL - 24
SP - 258
EP - 269
JO - Placenta
JF - Placenta
SN - 0143-4004
IS - 2-3
ER -