Adenovirus-delivered DKK3/WNT4 and steroidogenesis in primary cultures of adrenocortical cells
Adenovirus-delivered DKK3/WNT4 and steroidogenesis in primary cultures of adrenocortical cells
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DOI：
10.1055/s-2006-950500
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发表时间：
2006-09-01
影响因子：
2.2
通讯作者：
Hornsby, P. J.
中科院分区：
文献类型：
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作者：
Chen, M.;Hornsby, P. J.
Wnt家族分子Dickkopf-3(DKK3)和Wnt4在肾小球带中的浓度高于肾上腺皮质的其他部分。为了研究这些蛋白对肾上腺皮质细胞功能的直接影响，我们创造了编码人DKK3和WNT4的腺病毒。当添加到培养的人肾上腺皮质细胞中时，DKK3单独或与环磷酸腺苷一起抑制醛固酮和皮质醇的生物合成。WNT4单独添加时增加类固醇合成，但在环磷酸腺苷存在下减少类固醇合成。而携带GFP基因的腺病毒对照组则没有作用。从培养的细胞中提取RNA，用实时荧光定量聚合酶链式反应(Real-time PCR)进行检测。Cyp11a1(胆固醇侧链裂解酶)、HSD3B2(3β-羟基类固醇脱氢酶II)、CYP17(17α-羟基酶)、CYP21(21-羟基酶)和CYP11B1(11β-羟基酶)的mRNAs均被环磷酸腺苷所增加，而醛固酮合成酶则不受影响。DKK3减少cAMP刺激的细胞色素P17，Wnt4在没有环磷酸腺苷的情况下增加细胞色素P17和细胞色素P21。DKK3和WNT4均使细胞色素P11B2水平升高。这些数据表明，这些Wnt信号分子对肾上腺皮质细胞的类固醇合成具有多方面的作用，包括对整个类固醇合成(醛固酮和皮质醇的生物合成)的影响，以及对类固醇生成酶mRNA水平的不同影响。DKK3和WNT4在肾小球中的共同定位及其对细胞色素P11B2的刺激暗示了对肾小球特异性功能的作用。
The Wnt family molecules Dickkopf-3 (DKK3) and WNT4 are present at higher concentrations in the zona glomerulosa than in the rest of the adrenal cortex. In order to study direct effects of these proteins on adrenocortical cell function, we created adenoviruses encoding human DKK3 and WNT4. When added to cultured human adrenocortical cells, DKK3 inhibited aldosterone and cortisol biosynthesis, either alone or together with cyclic AMP. WNT4 increased steroidogenesis when added alone but decreased it in the presence of cyclic AMP. A control adenovirus encoding GFP had no effect. RNA was prepared from cultured cells and was assayed by real-time PCR. CYP11A1 (cholesterol side-chain cleavage enzyme), HSD3B2 (3 beta-hydroxysteroid dehydrogenase type II), CYP17 (17 alpha-hydroxylase), CYP21 (21-hydroxylase) and CYP11B1 (11 beta-hydroxylase) mRNAs were all increased by cyclic AMP, whereas CYP11B2 (aldosterone synthase) was unaffected. DKK3 decreased cyclic AMP-stimulated CYP17 WNT4 increased both CYP17 and CYP21 in the absence of cyclic AMP. Both DKK3 and WNT4 increased the level of CYP11B2. These data show that these Wnt signaling molecules have multiple actions on steroidogenesis in adrenocortical cells, including effects on overall steroidogenesis (aldosterone and cortisol biosynthesis) and distinct effects on steroidogenic enzyme mRNA levels. The co-localization of DKK3 and WNT4 in the glomerulosa and their stimulation of CYP11B2 imply an action on glomerulosa-specific function.