Koyama K, Krozowski Z
Laboratory of Molecular Hypertension, Baker Medical Research Institute, P.O. Box 6492, St. Kilda Rd. Central, Melbourne 8008, Australia.
Mol Cell Endocrinol. 2001 Oct 25;183(1-2):165-70. doi: 10.1016/s0303-7207(01)00548-2.
An important determinant of the potency of steroid hormones is the presence of activating and inactivating enzymes in target cells. The 11 beta-hydroxysteroid dehydrogenase type 1 and type 2 enzymes (11 beta HSD1 and 11 beta HSD2) modulate glucocorticoid action and may be important in regulating cellular growth. In the present study we examined 11 beta-hydroxysteroid dehydrogenase in Ishikawa endometrial cancer cells to see if modulation of enzyme activity could potentiate the antiproliferative effects of glucocorticoids. Ishikawa cells contain an NAD dependent enzyme migrating at 41 kDa on Western blots, consistent with the presence of the glucocorticoid-inactivating enzyme 11 beta HSD2, while the NADP dependent 11 beta HSD1 is barely detectable. Given that glucocorticoids decrease cellular proliferation we asked whether inhibition of 11 beta HSD2 could further enhance this effect. Cultivation of cells in the presence of 1 microM cortisol resulted in an elevation of 11 beta HSD2 and this was associated with a decrease in cell number. Enzyme activity and cell proliferation showed a biphasic response to the synthetic anti-progestin and anti-glucocorticoid RU38486, with < or =10 nM exerting agonistic effects and > or =100 nM producing antagonist effects in the presence of 1 microM cortisol. Inhibition of 11 beta HSD2 activity by glycyrrhetinic acid did not enhance the anti-proliferative effects of 1 microM cortisol, but the inhibitor showed significant antiproliferative activity in the absence of added glucocorticoid, consistent with protection of the low levels of glucocorticoids present in culture medium. Interestingly, the commonly used 11 beta HSD inhibitor, Carbenoxolone, did not block 11 beta HSD2 activity in whole Ishikawa cells, and there was no effect on cell proliferation, however, complete inhibition of 11 beta HSD2 was achieved in cellular homogenates suggesting that a barrier exists to entry of the inhibitor into intact cells. This study suggests that inhibition of 11 beta HSD2 activity can enhance the antiproliferative effects of low, but not high concentrations of glucocorticoids, and that beneficial effects may be attained in vivo at the nadir of diurnal glucocorticoid levels.
类固醇激素效力的一个重要决定因素是靶细胞中存在激活酶和失活酶。11β-羟基类固醇脱氢酶1型和2型酶（11βHSD1和11βHSD2）调节糖皮质激素的作用，可能在调节细胞生长中起重要作用。在本研究中，我们检测了石川子宫内膜癌细胞中的11β-羟基类固醇脱氢酶，以观察酶活性的调节是否能增强糖皮质激素的抗增殖作用。石川细胞含有一种NAD依赖性酶，在蛋白质印迹上迁移至41 kDa，这与糖皮质激素失活酶11βHSD2的存在一致，而NADP依赖性的11βHSD1几乎检测不到。鉴于糖皮质激素会降低细胞增殖，我们询问抑制11βHSD2是否能进一步增强这种作用。在1μM皮质醇存在下培养细胞导致11βHSD2升高，这与细胞数量减少有关。酶活性和细胞增殖对合成抗孕激素和抗糖皮质激素RU38486表现出双相反应，在1μM皮质醇存在下，≤10 nM发挥激动作用，≥100 nM产生拮抗作用。甘草次酸抑制11βHSD2活性并未增强1μM皮质醇的抗增殖作用，但该抑制剂在未添加糖皮质激素的情况下显示出显著的抗增殖活性，这与保护培养基中存在的低水平糖皮质激素一致。有趣的是，常用的11βHSD抑制剂甘珀酸在完整的石川细胞中并未阻断11βHSD2活性，对细胞增殖也没有影响，然而，在细胞匀浆中实现了11βHSD2的完全抑制，这表明抑制剂进入完整细胞存在障碍。这项研究表明，抑制11βHSD2活性可以增强低浓度而非高浓度糖皮质激素的抗增殖作用，并且在昼夜糖皮质激素水平最低点时，体内可能会获得有益效果。
