雌激素在小儿血管瘤增生机制中的作用及临床意义
摘要
[目的] 探讨小儿血管瘤血管生成的促发机制，阐明雌激素在小儿血管瘤增生机制中的促血管生成作用，为抗雌激素药物治疗小儿血管瘤提供新的理论依据。
[方法] 应用免疫组织化学SP法检测42例血管瘤和17例血管畸形标本雌激素受体(ER)、血管内皮生长因子(VEGF)、碱性成纤维细胞生长因子(bFGF)的表达情况，分析其相关性；应用纤维蛋白凝胶(fibrin gel)三维培养体系建立血管瘤血管生成体外培养模型，以雌激素及抗雌激素受体阻滞剂三苯氧胺分组(分空白对照组、雌激素组、三苯氧胺组、雌激素+三苯氧胺组四组)干预血管生成过程，比较各时间点各组间新生血管区面积的显著性差异。
[结果]
1、血管瘤组的ER、VEGF、bFGF平均标记指数均高于血管畸形及正常皮肤组，血管瘤组中增生期亚组三指标标记指数均高于退化期亚组(P＜0.01)。血管瘤中ER与VEGF、bFGF具强正相关性，其相关系数分别为：0.91、0.83，而血管畸形组中该相关性较弱，相关系数分别为：0.20、0.58(P＜0.05)。
2、血管瘤血管生成体外培养模型中小儿血管瘤组织培养2-3天后芽生出细小血管，后呈枝芽状生长，至第8-9天长成树枝状血管，分叉交叠，之后血管生长渐缓停滞。该模型中所取标本行病理观察证实为增生期血管瘤组织。组织周边新生树枝状结构经石蜡切片HE染色、血管内皮细胞特异性标志物
第姗因子相关抗原检测及电镜观察鉴定为血管。
3、培养后第3、6、9天雌激素组组织块新生血管区面积均显著大于空白对照
组，雌激素+三苯氧胺组及三苯氧胺组新生血管区面积均显著小于雌激素
组及空白对照组(P<0.05)。
〔结论〕
1、结合免疫组化实验及体外培养实验可以认为雌激素通过与ER结合能促进小
儿血管瘤血管生成，从而促进血管瘤增殖。其机制可能为雌激素与ER结合
后上调VEGF、bFGF的表达，并/或与VEGF、bFGF协同促进血管生成，该过
程能被三苯氧胺所阻滞。
2、血管瘤血管生成三维体外培养模型的建立，为研究血管瘤增殖退化过程中血
管生成的调节机制提供良好的模型，并有助于抗雌激素及抗血管生成药物的
研究，为药物治疗小儿血管瘤提供新的途径。
Objective: To study the promotion mechanism of angiogenesis of children hemangioma, to clarify the angiogenesis function of estrogen on the proliferative mechanism of children hemangioma, and also to provide the new theoretical evidence for ant-estrogen drugs cured children hemangioma.
Mothod: Expression and relativity of estrogen receptor(ER), VEGF and bFGF were detected with SP immunohistochemical method in 42 specimens of hemangioma and 17 specimens of vascular malformation. A fragment of hemangioma biopsy was embedded in fibrin gel, and a in vitro human model of angiogenesis of hemangioma was founded. The angiogenesis of hemangioma fragments of each group was interfered by estrogen and tomaxifen, and the dimension of newborn tubule area in the 3rd, 6th, 9th day after culture was calculated to compare statistically significant differences among the groups.
Result:
1. The label index (LI) of ER, VEGF and bFGF in the hemangioma were
significantly higher than those in the vascular malformation
(p<0.01) and in the hemangioma the LI of ER. VEGF and bFGF in
proliferative stage were higher than those in involuting
stage(p<0.01). There were significantly positive correlations between the LI of ER and VEFG, bFGF strong in hemangioma and weak
in the vascular malformantion(p<0. 05).
2. In the model of angiogenesis of hemangioma, microvessels grew out from the tissue fragments in 2nd to 3th day after culture, and in 8th to 9th day a complex network of microvessels come to being, the tending to inactivity. The complex network around the tissue fragments was confirmed to he vascular by immunohistochemistry experimentation and electron microscopic observation.
3. In the 3rd, 6th and 9th day after culture the dimension of newborn tubule area of the group of estrogen were significantly larger than those of the group of control. Those of the group of estrogen +tomaxifen and the group of tomaxifen were significant smaller than those of the group of estrogen and the group of control. (p<0. 05)
Conclusion:
1. Through the experimentation of immunohistochemistry and the study of culture in vitro it is demonstrated that estrogen stimulates angiogenesis of children hemangioma. The mechanism is possibly that estrogen bound to estrogen receptor leads to raise the expression of VEGF and bFGF, and/or cooperate with VEGF and bFGF to stimulate angiogenesis. Tomaxifen can hold back this process.
2. The model of angiogenesis of hemangioma provides a good model of researching the regulation mechanism of angiogenesis of hemangioma, and help to study the drugs of ant-estrogen and ant-angiogenesis to
provide a new approach for medication for children hemangioma.
[方法] 应用免疫组织化学SP法检测42例血管瘤和17例血管畸形标本雌激素受体(ER)、血管内皮生长因子(VEGF)、碱性成纤维细胞生长因子(bFGF)的表达情况，分析其相关性；应用纤维蛋白凝胶(fibrin gel)三维培养体系建立血管瘤血管生成体外培养模型，以雌激素及抗雌激素受体阻滞剂三苯氧胺分组(分空白对照组、雌激素组、三苯氧胺组、雌激素+三苯氧胺组四组)干预血管生成过程，比较各时间点各组间新生血管区面积的显著性差异。
[结果]
1、血管瘤组的ER、VEGF、bFGF平均标记指数均高于血管畸形及正常皮肤组，血管瘤组中增生期亚组三指标标记指数均高于退化期亚组(P＜0.01)。血管瘤中ER与VEGF、bFGF具强正相关性，其相关系数分别为：0.91、0.83，而血管畸形组中该相关性较弱，相关系数分别为：0.20、0.58(P＜0.05)。
2、血管瘤血管生成体外培养模型中小儿血管瘤组织培养2-3天后芽生出细小血管，后呈枝芽状生长，至第8-9天长成树枝状血管，分叉交叠，之后血管生长渐缓停滞。该模型中所取标本行病理观察证实为增生期血管瘤组织。组织周边新生树枝状结构经石蜡切片HE染色、血管内皮细胞特异性标志物
第姗因子相关抗原检测及电镜观察鉴定为血管。
3、培养后第3、6、9天雌激素组组织块新生血管区面积均显著大于空白对照
组，雌激素+三苯氧胺组及三苯氧胺组新生血管区面积均显著小于雌激素
组及空白对照组(P<0.05)。
〔结论〕
1、结合免疫组化实验及体外培养实验可以认为雌激素通过与ER结合能促进小
儿血管瘤血管生成，从而促进血管瘤增殖。其机制可能为雌激素与ER结合
后上调VEGF、bFGF的表达，并/或与VEGF、bFGF协同促进血管生成，该过
程能被三苯氧胺所阻滞。
2、血管瘤血管生成三维体外培养模型的建立，为研究血管瘤增殖退化过程中血
管生成的调节机制提供良好的模型，并有助于抗雌激素及抗血管生成药物的
研究，为药物治疗小儿血管瘤提供新的途径。
Objective: To study the promotion mechanism of angiogenesis of children hemangioma, to clarify the angiogenesis function of estrogen on the proliferative mechanism of children hemangioma, and also to provide the new theoretical evidence for ant-estrogen drugs cured children hemangioma.
Mothod: Expression and relativity of estrogen receptor(ER), VEGF and bFGF were detected with SP immunohistochemical method in 42 specimens of hemangioma and 17 specimens of vascular malformation. A fragment of hemangioma biopsy was embedded in fibrin gel, and a in vitro human model of angiogenesis of hemangioma was founded. The angiogenesis of hemangioma fragments of each group was interfered by estrogen and tomaxifen, and the dimension of newborn tubule area in the 3rd, 6th, 9th day after culture was calculated to compare statistically significant differences among the groups.
Result:
1. The label index (LI) of ER, VEGF and bFGF in the hemangioma were
significantly higher than those in the vascular malformation
(p<0.01) and in the hemangioma the LI of ER. VEGF and bFGF in
proliferative stage were higher than those in involuting
stage(p<0.01). There were significantly positive correlations between the LI of ER and VEFG, bFGF strong in hemangioma and weak
in the vascular malformantion(p<0. 05).
2. In the model of angiogenesis of hemangioma, microvessels grew out from the tissue fragments in 2nd to 3th day after culture, and in 8th to 9th day a complex network of microvessels come to being, the tending to inactivity. The complex network around the tissue fragments was confirmed to he vascular by immunohistochemistry experimentation and electron microscopic observation.
3. In the 3rd, 6th and 9th day after culture the dimension of newborn tubule area of the group of estrogen were significantly larger than those of the group of control. Those of the group of estrogen +tomaxifen and the group of tomaxifen were significant smaller than those of the group of estrogen and the group of control. (p<0. 05)
Conclusion:
1. Through the experimentation of immunohistochemistry and the study of culture in vitro it is demonstrated that estrogen stimulates angiogenesis of children hemangioma. The mechanism is possibly that estrogen bound to estrogen receptor leads to raise the expression of VEGF and bFGF, and/or cooperate with VEGF and bFGF to stimulate angiogenesis. Tomaxifen can hold back this process.
2. The model of angiogenesis of hemangioma provides a good model of researching the regulation mechanism of angiogenesis of hemangioma, and help to study the drugs of ant-estrogen and ant-angiogenesis to
provide a new approach for medication for children hemangioma.
引文
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7 Mulliken JB. et al. In vitro tharacteristics of endothelium from hemangiomes and vascular malformations. Surgery(St. louis), 1982, 92: 348-353
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9 Floman J. et al. Clinical applications of reseach on angiogenesis. N Engl J Med, 1995, 333: 1757
10 Vailhe B. et al. In vitro models of vasculogenesis and angiogenesis. Lab Invest, 2001, 81(4): 439-452
11 初同伟．血管内皮细胞生长因子的作用及调节．第三军医大学学报，2000，22(7)：705—707
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angiogenesis. J Surg Res, 1997, 67(2): 147-154
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2 吕麟亚，邓辉胜．儿童血管瘤发病机制研究进展．医学综述，2000，6(1)：26—27
3 Boye E. et al. Clonality and altered behavior of endothelial cells form hemangiomas. J Clin Invest, 2001, 107 (6): 745-752
4 胡琼华，王炜．血管瘤及血管畸形的增殖机制及治疗进展．实用美容整形外科杂志，2000，11(2)：95—98
5 徐泉，李恭才．血管瘤的研究现状．中级医刊，1996，3l(3)：12—14
6 Mulliken JB et al. hemangiomas and malformationsin infants and childien: a classification based on endothelial characteristics. Plas Reconstr Surg, 1982, 63: 412-420
7 Mulliken JB. et al. In vitro tharacteristics of endothelium from hemangiomes and vascular malformations. Surgery(St. louis), 1982, 92: 348-353
8 Takahashi K. et al. Cellular markers that distinguish the phase of hemangioma during infancy and childhood. J Clin Invest, 1994, 93: 2357-2364
9 Floman J. et al. Clinical applications of reseach on angiogenesis. N Engl J Med, 1995, 333: 1757
10 Vailhe B. et al. In vitro models of vasculogenesis and angiogenesis. Lab Invest, 2001, 81(4): 439-452
11 初同伟．血管内皮细胞生长因子的作用及调节．第三军医大学学报，2000，22(7)：705—707
12 郭伟，王莉莉．血管内皮细胞生长因子的研究进展．新乡医学院学报，2001，18(1)：69—71
13 Zhang QX. et al. Vascular endothelial growth factor is the major angiogenic factor in omentum mechanism of the omentum-mediated
angiogenesis. J Surg Res, 1997, 67(2): 147-154
14 Ryuto M. et at. Induction of vascular endothelial growth factor by tumor necrosis factor alpha in human glioma cells, possible roles of SP-1. J Biol Chem, 1996, 271(45): 28220-28225
15 Collins PD. et al. Characterization of the increase invascular permeability factor in vivo. Br J Pharmacol, 1993, 109(1): 195-199
16 Chang J. et al. Proliferative hemangiomas: analysis of cytokine gene expression and angiogenesis. Plast Reconstr Surg, 1999, 103(1): 1-9
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