| 研究実績の概要 |
We are hypothesizing that estrogen receptor β (ERβ) may mediates anti-inflammatory effect in colitis through epigenetic regulation. First of all, immunohistochemistry revealed the ERβ, but not ERα, was expressed in duodenum and colon, but not in jejunum and ileum. Similar findings were obtained by western blotting and RT-PCR, that indicates ERβ dominance in intestinal tissue. In addition, southwestern histochemistry was detected highly similar co-localization of ERE-binding proteins with ERβ indicated that ERβ is functionally active in intestinal epithelium. Moreover, to investigate epigenetic treatment approach is useful in colitis model, we examined the effects of the histone deacetylase (HDAC) inhibitor, suberoylanilide hydroxamic acid (SAHA) in 1.5% dextran sulfate sodium (DSS) induced colitis model. Pro-inflammatory cytokines IL-6, TNF-α, and chemokine Ccl2 gene expression peaked on day 5 in DSS-treated mouse colon, whereas SAHA treatment significantly decreased pro-inflammatory gene expression. In addition, the localization of CD11b showed that migratory inflammatory cells were dramatically decreased by SAHA treatment compared to DSS-treated mouse colon. Thus, we conclude that the HDAC inhibitor, SAHA, attenuates inflammatory changes in DSS-induced colitis by suppressing local secretion of pro-inflammatory cytokines and chemokines. Altogether these results indicate that epigenetic treatment approach could be useful for colitis. In the next, we will examine the epigenetic regulation of ERβ in 1.5% DSS induced colitis model using WT and ERβ knockout mice.
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| 現在までの達成度 (区分) |
現在までの達成度 (区分)
3: やや遅れている
理由
To study the role of estrogen signaling in mouse intestinal tract, we examined the expression of ERα and ERβ in male and female age-matched mouse. In intestinal epithelium, ERβ was expressed in duodenum and colon, but not in jejunum and ileum. However, ERα was not detected throughout intestinal epithelium. To examine the transcriptional activity of ERβ, we examined southwestern histochemistry and results revealed that the expression of ERE-binding proteins pattern was highly similar to that of ERβ. Moreover, to optimize colitis model in mouse, 1-3% DSS was given with drinking water to C57BL/6J mice. Based on the results, 1.5% DSS was evaluated as optimal concentration to induce colitis in mouse. In addition, the effects of SAHA was examined in colitis model. Pro-inflammatory cytokines IL-6, TNF-α, and chemokine Ccl2 gene expressions were increased in DSS-treated mouse colon, whereas SAHA treatment significantly decreased pro-inflammatory gene expression.
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| 今後の研究の推進方策 |
As for next experiment, to study estrogen effect on epigenetic regulation, 17β-estradiol will be treated with and without ER inhibitor. Direct involvement of ERβ in the epigenetic regulation will be investigated in ERβKO mouse colon. DSS-induced colitis model will be generated using ERβKO mouse and detail epigenetic analysis will be performed. Moreover, Caco-2 and T84 colonic epithelial cell lines will be used for analysis of epigenetic regulation and its involvement of estrogen and ERβ.
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