Division of Endocrinology and Metabolism, Rostock University Medical Center, Ernst-Heydemann-Straße 6, 18057, Rostock, Germany.
Department of Urology, Rostock University, Rostock, Germany.
Cell Tissue Res. 2021 May;384(2):325-331. doi: 10.1007/s00441-021-03432-9. Epub 2021 Mar 2.
It still remains unclear how the functional organisation of the adrenal cortex arises. One aim of this study was to create a setup which allows for the establishment of a concentration gradient in vitro. This was achieved by a continuous flow of medium through the culture flask which caused differences in glucose and cortisol concentrations as well as in pH values between the sites of inflow and outflow of medium. Using real-time polymerase chain reaction, we found that a continuous supply of 1 ml medium per hour significantly increased the expression of MC2R, CYP11B1 and CYP17A1 genes of NCI-H295R cells in the distal area of the flask as compared with the proximal part. The expression of the AT1R showed a reverse regulation. The addition of dexamethasone to the medium led to an increase in gene expression of MC2R while AT1R was downregulated. Moreover, we detected a higher expression of CYP11B2 and a decreased expression of CYP11B1 when endothelial cell-conditioned medium (ECCM) was added to the inflow. Our experiments show that a directed medium delivery system creates different gradients and affects the functional differentiation of the NCI-H295R cells. Also, our results emphasise that products of endothelial cells have additional effects on the differentiation of the cultured adrenal cortical cells. Our results are in support that the regulation of the adrenal zonation is possible through different concentration gradients.
肾上腺皮质的功能组织是如何产生的仍不清楚。本研究的目的之一是建立一种体外建立浓度梯度的方法。通过培养基在培养瓶中的连续流动来实现这一点，这导致了流入和流出培养基的部位之间的葡萄糖和皮质醇浓度以及 pH 值的差异。使用实时聚合酶链反应，我们发现与瓶的近端部分相比，每小时连续供应 1 毫升培养基可显著增加 NCI-H295R 细胞在远端的 MC2R、CYP11B1 和 CYP17A1 基因的表达。AT1R 的表达呈现相反的调节。向培养基中添加地塞米松会导致 MC2R 的基因表达增加，而 AT1R 则下调。此外，当将内皮细胞条件培养基 (ECCM) 添加到流入物中时，我们检测到 CYP11B2 的表达增加，而 CYP11B1 的表达减少。我们的实验表明，定向的培养基输送系统会产生不同的梯度，并影响 NCI-H295R 细胞的功能分化。此外，我们的结果强调了内皮细胞产物对培养的肾上腺皮质细胞分化的额外影响。我们的结果支持通过不同的浓度梯度来调节肾上腺分带的观点。
