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ORIGINAL RESEARCH ARTICLE

Establishment of a Primary Culture Model of Mouse Uterine and Vaginal Stroma for Studying In Vitro Estrogen Effects

Keiko Inada^*, Shinji Hayashi^*, Taisen Iguchi^, and Tomomi Sato^*,1

^* Graduate School of Integrated Science, Yokohama City University, Yokohama 236-0027, Japan; The Graduate University for Advanced Studies and Okazaki Institute for Integrative Bioscience, National Institutes of Natural Sciences, Okazaki 444-8787, Japan; and CREST, Japan Science and Technology Corporation, Kawaguchi 332-0012, Japan

To whom requests for reprints should be addressed at ¹ Graduate School of Integrated Science, Yokohama City University, 22–2 Seto, Kanazawa-ku, Yokohama 236-0027, Japan. E-mail: tomomi{at}yokohama-cu.ac.jp

Effects of 17ß-estradiol (E2) on uterine and vaginal epithelial cell proliferation could be mediated by stromal cell–derived paracrine factors. To study the epithelial-stromal interactions in mice, an in vitro model of uterine and vaginal stromal cells of immature mice is essential. Therefore, we established a primary culture model of stromal cells both from uterus and vagina and examined the effect of E2 on proliferation of cultured stromal cells. We found that E2 stimulated proliferation of stromal cells from both organs in vitro, showing an increase in the number of cells and the percentage of 5-bromo-2'-deoxyuridine (BrdU)–labeled cells. Interestingly, vaginal stromal cells responded to lower E2 than uterine stromal cells in proliferation (10^–12 M vs. 10^–8 M) and BrdU labeling (10^–14 – 10^–10 M vs. 10^–10 – 10^–6 M). To examine the effect of E2 in vivo, cells were grafted into the subrenal capsule of the host mice and grown for 2 weeks. The BrdU labeling in cultured stromal cells was increased by E2 in vivo. To examine the effect of cultured stromal cells on epithelial cell proliferation, uterine and vaginal epithelium of adult mice were separated, recombined with the cultured stromal cells, and grafted under the renal capsule of hosts for 3 weeks. Epithelial cells recombined with cultured stromal cells showed simple columnar morphology in uterine grafts and stratified and keratinized morphology in vaginal grafts under the influence of the hormonal environment of the hosts. The BrdU labeling in epithelial cells was increased by E2, suggesting that cultured stromal cells can stimulate epithelial cell proliferation. In conclusion, we established a primary culture model of uterine and vaginal stromal cells, which can be mitogenically stimulated by E2 in vitro and in vivo after being grafted under the renal capsule. This culture system will be useful for investigating the underlying molecular mechanisms of uterine and vaginal epithelial-stromal interactions.

Key Words: uterus • vagina • stromal cells • estrogen