In situ estrogen synthesized by aromatase P450 in uterine leiomyoma cells promotes cell growth probably via an autocrine/intracrine mechanism.
In situ estrogen synthesized by aromatase P450 in uterine leiomyoma cells promotes cell growth probably via an autocrine/intracrine mechanism.
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DOI：
10.1210/endo.141.10.7719
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发表时间：
2000-10
期刊：
影响因子：
4.8
通讯作者：
Hiroshi Sumitani;Makio Shozu;T. Segawa;K. Murakami;Hui-Juan Yang;Keiko Shimada;M. Inoue
中科院分区：
文献类型：
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作者：
Hiroshi Sumitani;Makio Shozu;T. Segawa;K. Murakami;Hui-Juan Yang;Keiko Shimada;M. Inoue
在本研究中，我们详细描述了平滑肌瘤中芳香酶 P450 的表达，以确定原位雌激素在平滑肌瘤生长优势中的作用。通过定量 RT-PCR 测定，平滑肌瘤中的芳香酶 P450 转录物水平显着高于相应的子宫肌层。蛋白质印迹分析也证实了平滑肌瘤中芳香酶 P450 的过度表达。免疫反应性芳香酶的估计大小为 58 kDa，与胎盘中的相似。为了鉴定在平滑肌瘤中表达芳香酶 P450 的细胞类型，使用多克隆抗体对组织学标本进行芳香酶 P450 染色。在平滑肌瘤细胞的细胞质中检测到强烈的免疫反应性，而周围的正常子宫肌层则显示弱或阴性染色。从平滑肌瘤获得的培养物中的平滑肌样细胞，肌动蛋白 D 纤维呈阳性，细胞质中具有芳香酶的免疫反应颗粒。佛波醇肉豆蔻酸酯乙酸酯和地塞米松加白细胞介素 1β 有效刺激雄激素向雌激素的转化，并被选择性芳香酶 P450 抑制剂（法屈唑和 TZA-2209）完全消除，但不能被 5α 还原酶抑制剂（非那雄胺和氟他胺）消除。在地塞米松和白介素-1β存在下，雄烯二酮的表观 Km 为 3 nM，对应于育龄妇女的血浆雄烯二酮浓度。为了确定内源性芳香酶 P450 是否在促进平滑肌瘤细胞生长中发挥作用，我们使用 WST-1 测定评估了用不同浓度的雌激素和雄激素处理的平滑肌样细胞的细胞生长。睾酮（10（-8）和10（-7）M）和雄烯二酮（10（-8）和10（-7）M）刺激从平滑肌瘤获得的平滑肌样细胞的生长，其程度与雌二醇（10（-10）-10（-7）M）相同，而二氢睾酮（10（-11）-10（-8）M）则不然。与法屈唑共同治疗再次消除了睾酮对细胞生长的刺激作用。睾酮(10(-8)M)处理的平滑肌样细胞培养基中雌二醇的水平为10(-11)M，比促进细胞生长所需的最低雌二醇浓度(10(-10)M)低1个数量级。这表明平滑肌瘤中合成的雌二醇通过自分泌/内分泌机制促进其生长。我们得出结论，平滑肌瘤的子宫肌细胞过度表达芳香酶 P450，并且能够合成足够的雌激素以加速其自身细胞生长。芳香酶 P450 的过度表达可能通过自分泌/内分泌机制在平滑肌瘤组织相对于周围子宫肌层的生长优势中发挥作用。
In the present study we characterized in detail the expression of aromatase P450 in leiomyomas to determine the role of in situ estrogen in the growth advantage of leiomyomas. The levels of aromatase P450 transcripts were determined by quantitative RT-PCR to be significantly higher in leiomyomas than in corresponding myometrium. The overexpression of aromatase P450 in leiomyomas was also confirmed by Western blot analysis. The estimated size of immunoreactive aromatase was 58 kDa, similar to that in placenta. To identify a cell type that express aromatase P450 in leiomyomas, histological specimens were stained for aromatase P450 using a polyclonal antibody. Strong immunoreactivity was detected in the cytoplasm of leiomyoma cells, whereas surrounding normal myometrium displayed weak or negative staining. Smooth muscle-like cells in culture obtained from leiomyomas, positive for actin D fiber, possessed immunoreactive granules of aromatase in the cytoplasm. Conversion of androgen to estrogen was effectively stimulated by phorbol myristate acetate and dexamethasone plus interleukin-1beta and was completely abolished by selective inhibitors of aromatase P450 (fadrozole and TZA-2209), but not by inhibitors of 5alpha-reductase (finasteride and flutamide). The apparent Km of androstenedione was 3 nM in the presence of dexamethasone and interleukin-1beta, corresponding to the plasma concentration of androstenedione in women of reproductive age. To determine whether endogenous aromatase P450 plays a role in the growth promotion of leiomyoma cells, we evaluated the cell growth of smooth muscle-like cells treated with various concentrations of estrogen and androgen using a WST-1 assay. Treatment with testosterone (10(-8) and 10(-7) M) and androstenedione (10(-8) and 10(-7) M) stimulated the growth of smooth muscle-like cells obtained from leiomyomas to the same extent as estradiol (10(-10)-10(-7) M), whereas dihydrotestosterone (10(-11)-10(-8) M) did not. The stimulatory effect of testosterone on cell growth was again abolished by cotreatment with fadrozole. The level of estradiol in the medium of testosterone (10(-8) M)-treated smooth muscle-like cells was 10(-11) M, which was 1 order lower than the minimum concentration of estradiol necessary to promote cell growth (10(-10) M). This indicates that estradiol synthesized in leiomyomas promotes their growth via an autocrine/intracrine mechanism. We conclude that myometrial cells of leiomyomas overexpress aromatase P450 and are able to synthesize sufficient estrogen to accelerate their own cell growth. Overexpression of aromatase P450 may play a role in the growth advantage of leiomyoma tissue over surrounding myometrium via an autocrine/intracrine mechanism.