2007 Fiscal Year Final Research Report Summary
Development of in vitro model for implantation using the endometrial spheroid
| Project/Area Number |
18580282
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Applied animal science
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| Research Institution | Kyushu University |
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Principal Investigator |
YAMAUCHI Nobuhiko Kyushu University, Faculty of Agriculture, Associate Professor (00363325)
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| Co-Investigator(Kenkyū-buntansha) |
HATTORI Massaki Kyushu University, Faculty of Agriculture, Professor (60175536)
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| Project Period (FY) |
2006 – 2007
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| Keywords | spheroid / embryo / rat / three-dimensional culture / implantation / MMP |
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| Research Abstract |
The endometrium is one of the most complex tissues; it undergoes dynamic changes in response to implantation and pregnancy processes. An in vitro model may provide a tool for clarifying the complex implantation process. However, there is no suitable in vitro model for the investigation of endomotrial functions. The spheroid has been utilized in cell biology research because it appears to mimic morphology and physiology of cells in living tissues and organs, which is unlike conventional monolayer culture. Multicellular spheroids composed of normal adult cells may provide a more useful model for the study of endometrium. The purpose of the present study is to develop the spheroid composed of rat endometrial stromal cells(RES) as an in vitro model for analysis of the endometrial functions.
The RES were prepared from rat endometrium at day 5 of pregnancy. Spheroids were generated using salmon aterocollagen(SAC). The cells were plated on 12 well SAC gels(Imoto Suisan Co.) and cultured in the
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culture medium(DMEM/F12) containing 10% foetal bovine serum. After the cells reached confluence at day 7 in culture, SAC gels were digested by collagenase to promote the detachment of RES cell sheets. Then the floating cell sheets were transferred to agarose-coated plates and cultured to form spheroids. The cell-sheet shrank and became an aggregated cell mass in a few days; it finally formed a round-shaped spheroid. Diameters of the spheroids were about 500μm at day 5 after detachment from SAC gels.
TUNEL examination of cell viability in the spheroid suggested no cell was apoptotic until 15 days after cell sheet detachment, TUNEL-positive cells appeared at 20 days. Additionally, no positive staining of PCNA was observed in the spheroids in contrast to the strong staining in the proliferating monolayer cultured cells. The results of gelatin zymography showed that both MMP-2 and -9 were produced in monolayer culture. However, after the detachment of the cell sheet, the production of both MMPs decreased immediately and could not be detected until 15 days after detachment.In vitro decidualization of the spheroids were induced by arachidonic acid(AA) treatment. RT-PCR analysis showed that typical marker genes for the decidualization, desmin and d/tPRP, were expressed in the spheroids after 2 days of AA treatment, but not in the control groups without treatment.
The results of the present study indicate that rat endometrial stromal cells are capable of being regenerated as a spheroid using SAC gels in vitro. Importantly, the present spheroid displays an endometrium-mimic feature in both the structural and functional similarities. The present method is simple and convenient, and therefore provides a new insight into the study of endometrial functions and implantation. Less
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[Journal Article] Expression of hedgehog family genes in the rat uterus during early pregnancy2008
Author(s)
K., Kubota, N., Yamauchi, K., Matsumoto, R., Watanabe, S., Oozono, S., Aramaki, C., Wood, T., Soh, M-A., Hattori
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Journal Title
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] Developmant of a rat endometrial spheroid as a model for the analysis of endometrial functions2008
Author(s)
K., Nishiraura, N., Yamauchi, R., Watanabe, K., Matsumoto, S., Oozono, K., Kubota, C., Wood, T., Soh, M-A., Hattori
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Journal Title
J Exp Zool Part A Ecol Genet Physiol (in press)
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] Expression of rat uterine serine proteinases homologous to mouse implantation serine proteinase 22008
Author(s)
S., Oozono, N., Yamauchi, K., Nishimura, K., Matsumoto, R., Watanabe, K., Kubota, S., Aramaki, F., Sato, C., Wood, T., Soh, M-A., Hattori
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Journal Title
J Exp Zool Part B Mol Dev Evol (in press)
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] In vitro decidualization of rat endometrial stromal cells2008
Author(s)
K., Matsumoto, N., Yamauchi, R., Watanabe, S., Oozono, K., Kubota, K., Nishimura, C., Wood, T., Soh, K., Kizaki, M-A., Hattori
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Journal Title
Cell Tissue Res (in press)
Description
「研究成果報告書概要(欧文)」より
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[Journal Article] Effects of tumor necrosis factor-alpha on cell proliferation, prostaglandins and matrix-metalloproteinases production in rat endometrial stromal cells cultured in vitro2007
Author(s)
T., Gamo, N., Yamauchi, K., Nishimura, R., Watanabe, K., Matsumoto, P-J., He, T., Soh, M-A., Hattori
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Journal Title
J Exp Zool Part A Ecol Genet Physiol 307
Pages: 699-707
Description
「研究成果報告書概要(欧文)」より
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[Presentation] Expression of three Hedgehog genes in rat uterus during early pregnancy2008
Author(s)
K., Kubota, S., Oozono, K., Matsumoto, R., Watanabe, T., Soh, N., Yamauchi, M-A., Hattori
Organizer
1st Word Congress on Reproductive Biology
Place of Presentation
Kona, Hawaii
Year and Date
20080000
Description
「研究成果報告書概要(欧文)」より
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[Presentation] In vitro decidualization of rat endometrial stromal cells2008
Author(s)
K., Matsumoto, R., Watanabe, S., Oozono, K., Kubota, K., Nishimura, T., Soh, N., Yamauchi, M-A., Hattori
Organizer
1st Word Congress on Reproductive Biology
Place of Presentation
Kona, Hawaii
Year and Date
20080000
Description
「研究成果報告書概要(欧文)」より
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[Presentation] Expression of matrix metalloproteinases in rat uterus during implantation phase2008
Author(s)
R., Watanabe, K., Matsumoto, S., Oozono, K., Kubota, K., Nishimura, T., Soh, N., Yamauchi, M-A., Hattori
Organizer
1st Word Congress on Reproductive Biology
Place of Presentation
Kona, Hawaii
Year and Date
20080000
Description
「研究成果報告書概要(欧文)」より
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[Presentation] Developmant of a rat endometrial spheroid as a model for the analysis of endometrial functions2007
Author(s)
K., Nishimura, S., Oozono, H., Urata, P-J., He, F., Sato, N., Yamauchi, M-A., Hattori
Organizer
33rd Annual Conference of the International Embryo Transfer Society
Place of Presentation
Kyoto, Japan
Year and Date
20070000
Description
「研究成果報告書概要(欧文)」より
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[Presentation] Development of spheroidal model using aterocollagen for the analysis of rat endometrial functions2006
Author(s)
K., Nishimura, S., Oozono, H., Urata, T., Soh, N., Yamauchi, M-A., Hattori
Organizer
12th Asian-Australasian Association of Animal Production Societies
Place of Presentation
Busan, Korea
Year and Date
20060000
Description
「研究成果報告書概要(欧文)」より
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[Presentation] The expression of implantation serine protease in the rat uterus2006
Author(s)
S., Oozono, H., Urata, K., Nishimura, P-J., He, T., Soh, N., Yamauchi, M-A., Hattori
Organizer
12th Asian-Australasian Association of Animal Production Societies
Place of Presentation
Busan, Korea
Year and Date
20060000
Description
「研究成果報告書概要(欧文)」より
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