Molecular mechanisms of dissociative glucocorticoid activity
Molecular mechanisms of dissociative glucocorticoid activity
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DOI：
10.1046/j.1365-2362.2000.0300s3006.x
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发表时间：
2000-12-01
影响因子：
5.5
通讯作者：
Schulte, HM
中科院分区：
文献类型：
--
作者：
Bamberger, CM;Schulte, HM
背景 糖皮质激素通过某些靶基因的反式激活和反式抑制来介导其对靶细胞的作用。虽然传统的糖皮质激素无法区分反式激活和反式抑制，但新的糖皮质激素应该能够消除这些作用，从而降低糖皮质激素在临床使用中出现不良副作用的可能性。在这项研究中，我们开发了一种新的实验系统，以在正常淋巴细胞中保留潜在选择性的糖皮质激素。 材料和方法 在用植物血凝素预处理后，使用电穿孔法转染正常淋巴细胞，其中 pGL3 荧光素酶报告载体受以下控制：（1）人 IL-2 启动子； (2)糖皮质激素反应元件(GRE)。测量对各种类固醇化合物的荧光素酶活性，包括潜在解离的糖皮质激素醋酸甲羟孕酮 (MPA)。结果 IL-2 启动子被佛波酯和离子霉素诱导 267.2 +/- 27.5 倍（平均值 +/- SD）。在这些细胞中，氢化可的松和地塞米松分别导致荧光素酶活性降低 22.9 +/- 3.6% 和 38.4 +/- 10%。在 GRE 控制下，氢化可的松刺激荧光素酶活性 6.4 +/- 0.50 倍，地塞米松刺激 8.2 +/- 0.4 倍。 MPA 诱导的 IL-2 启动子反式抑制为 73.3 +/- 7.2%，而 GRE 驱动构建体的反式激活为 2.4 +/- 0.4 倍。天然孕激素黄体酮对任一构建体均没有显着影响。结论这是第一个能够有效分析正常人淋巴细胞中糖皮质激素依赖性反式激活和反式抑制的系统。与传统糖皮质激素相比，MPA可称为解离性糖皮质激素，其反式阻抑/反式激活比为6.6（反式阻抑1.91/反式激活0.29），以地塞米松为标准（反式阻抑1/反式激活1）。我们得出结论，MPA是治疗自身免疫/炎症性疾病的一种非常有前景的物质。
Background Glucocorticoids mediate their effects on target cells via transactivation and transrepression of certain target genes. While conventional glucocorticoids do not distinguish between transactivation and transrepression, new glucocoticoids should be able to dissociate these effects, thus lowering the potential of unwanted side-effects of glucocorticoids in clinical use. In this study, we developed a new experimental system to rest potentially selective glucocorticoids in normal lymphocytes.Materials and methods Following pretreatment with phytohaemagglutinin, normal lymphocytes were transfected, using electroporation, with pGL3 luciferase reporter vectors under the control: (1) of the human IL-2 promoter; and (2) of a glucocorticoid response element (GRE). Luciferase activity was measured in response to various steroid compounds, including the potentially dissociative glucocorticoid medroxyprogesterone acetate (MPA).Results The IL-2 promoter was induced 267.2 +/- 27.5-fold (mean +/- SD) by phorbol ester and ionomycin. In these cells, hydrocortisone and dexamethasone caused a 22.9 +/- 3.6% and a 38.4 +/- 10% reduction in luciferase activity, respectively. Under GRE control, hydrocortisone stimulated luciferase activity 6.4 +/- 0.50-fold and dexamethasone 8.2 +/- 0.4-fold. MPA-induced transrepression was 73.3 +/- 7.2% for the IL-2 promoter, and transactivation was 2.4 +/- 0.4-fold with the GRE-driven construct. The natural progestin progesterone did not have significant effects on either construct.Conclusions This is the first system that allows efficient analysis of glucocorticoid-dependent transactivation and transrepression in normal human lymphocytes. Compared to conventional glucocorticoids, MPA can be reffered to as a dissociative glucocorticoid, its transrepression/transactivation ratio being 6.6 (transrepression 1.91/transactivation 0.29), with dexamethasone being the standard (transrepression 1/transactivation 1).We conclude chat MPA is a highly promising substance for the treatment of autoimmune/inflammatory diseases.