Mineralocorticoid and glucocorticoid receptor steroid binding and localization in colonic cells.
Mineralocorticoid and glucocorticoid receptor steroid binding and localization in colonic cells.
复制标题
盐皮质激素和糖皮质激素受体类固醇在结肠细胞中的结合和定位。
DOI：
10.1152/ajpcell.1994.266.3.c729
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发表时间：
1994
期刊：
影响因子：
--
通讯作者：
Bastl,CP
中科院分区：
文献类型：
--
作者：
Schulman,G;Robertson,NM;Elfenbein,IB;Eneanya,D;Litwack,G;Bastl,CP
在大鼠结肠上皮中，糖皮质激素和盐皮质激素通过与不同的受体结合并刺激不同的途径来调节钠转运。盐皮质激素（MR）和糖皮质激素（GR）受体在结肠钠吸收表面细胞和氯分泌隐窝细胞中的分布和细胞内定位尚不清楚。通过EDTA螯合和机械解离从大鼠远端结肠依次分离表面和隐窝细胞。通过台盼蓝排除和低比率的2 '，7'-双（2-羧乙基）-5（6）-羧基荧光素渗漏证实细胞活力。组织学检查，碱性磷酸酶活性，和[3 H]亮氨酸掺入率证实表面从隐窝细胞分离。[3 H]醛固酮和[3 H]曲安奈德结合的Scatchard分析表明，MR的数量从表面的7，228 +/-1，067减少到隐窝细胞中的2，299 +/- 434受体/细胞，而GR的数量从表面的20，857 +/-4，241增加到58，598 +/- 8，207个受体/细胞。MR的解离常数为2.8 +/- 0.4 nM，GR的解离常数为12 +/- 3 nM。使用特异性抗MR抗体hMRsN和抗GR抗体BuGR-2的间接免疫荧光表明，两种未配体的受体均为细胞质，并在激素结合后易位至细胞核。这些数据表明，表面和隐窝细胞是潜在的响应盐皮质激素和糖皮质激素和MR和GR需要激素的核转位。
In rat colon epithelium glucocorticoids and mineralocorticoids regulate Na transport by binding to distinct receptors and stimulating different pathways. The distribution and intracellular localization of mineralocorticoid (MR) and glucocorticoid (GR) receptors in colonic Na-absorbing surface cells and Cl-secreting crypt cells is unknown. Surface and crypt cells were sequentially isolated from rat distal colon by EDTA chelation and mechanical dissociation. Cell viability was confirmed by trypan blue exclusion and low rates of 2',7'-bis(2-carboxyethyl)-5(6)-carboxylfluorescein leak. Histologic examination, alkaline phosphatase activity, and rates of [3H]leucine incorporation confirmed separation of surface from crypt cells. Scatchard analysis of [3H]aldosterone and [3H]triamcinolone acetonide binding demonstrated that the number of MR decreased from 7,228 +/- 1,067 in surface to 2,299 +/- 434 receptors/cell in crypt cells, whereas the number of GR increased from 20,857 +/- 4,241 in surface to 58,598 +/- 8,207 receptors/cell in crypt cells. The dissociation constants were 2.8 +/- 0.4 nM for the MR and 12 +/- 3 nM for the GR. Indirect immunofluorescence using the specific anti-MR antibody hMRsN and the anti-GR antibody BuGR-2 demonstrated that both unliganded receptors were cytoplasmic and translocated to the nucleus after hormone binding. These data indicate that both surface and crypt cells are potentially responsive to mineralocorticoids and glucocorticoids and that both the MR and GR require hormone for nuclear translocation.