Clinical Research Unit, Centre of Internal Medicine, Justus Liebig University, Friedrichstrasse. 20/ Aulweg 123, 35392, Giessen, Germany.
Institute of Physiology, Justus Liebig University, Giessen, Germany.
Stem Cell Res Ther. 2019 Aug 6;10(1):240. doi: 10.1186/s13287-019-1330-x.
Adipose-derived mesenchymal stem cells (ADMSC) are non-haematopoietic, fibroblast-like multipotent progenitor cells. They have the potential for trilineage (adipocyte, chondrocyte and osteocyte) differentiation as well as differentiation into endocrine pancreatic progenitors. In diabetic or cancer therapy, somatostatin (SST) expression plays a vital role. Small molecules such as valproic acid (VPA) and micronutrients like vitamin D3 have differentiation potential in ADMSC. Therefore, the aim of this study was to investigate the role of vitamin D3 machinery and its metabolic enzymes in ADMSC. Furthermore, the reprogramming effect of vitamin D3 and VPA was evaluated on somatostatin expression in pancreatic lineage differentiation.
ADMSC were characterised based on their cell surface marker profile using flow cytometry. Specific adipogenic and osteogenic differentiation protocols were used in this study. Gene expression of several pluripotent, endodermal, pancreatic progenitor and pancreatic endocrine lineage markers were investigated in native ADMSC and after stimulation with different concentration of vitamin D3 for five consecutive days (0, 50, 100, 150 nM) and VPA (0.5, 1, 1.5, 2 mM) by real-time PCR. Furthermore, somatostatin expression was confirmed with ELISA and immunocytochemistry.
In ADMSC, the expression of somatostatin mRNA, the vitamin D receptor (VDR) and its metabolising enzymes 1 α-Hydroxylase, 24-Hydroxylase and 25-Hydroxylase were detected. Upon stimulation with vitamin D3, nuclear translocation of vitamin D receptor (VDR) was observed. Interestingly, the presence of vitamin D3 reduced the transcription of the somatostatin gene. By contrast, VPA treatment of cultivated ADMSC showed enhancing effect on somatostatin gene expression. No other pluripotent, endodermal, pancreatic progenitor or pancreatic endocrine lineage mRNA expression was modulated under the influence of vitamin D3 and VPA.
Human ADMSC carry the VDR. The vitamin D metabolising enzyme 25-Hydroxylase responded to the addition of vitamin D3. Moreover, our results demonstrate that somatostatin expression in ADMSC is constitutive, partially secreted and regulated by vitamin D3 and VPA.
脂肪间充质干细胞（ADMSC）是非造血的成纤维细胞样多能祖细胞。它们具有向三系（脂肪细胞、软骨细胞和成骨细胞）分化以及向内分泌胰腺祖细胞分化的潜能。在糖尿病或癌症治疗中，生长抑素（SST）的表达起着至关重要的作用。小分子如丙戊酸（VPA）和微量营养素如维生素 D3 具有 ADMSC 的分化潜力。因此，本研究的目的是研究维生素 D3 机制及其代谢酶在 ADMSC 中的作用。此外，还评估了维生素 D3 和 VPA 的重编程作用对胰腺谱系分化中生长抑素表达的影响。
使用流式细胞术根据其细胞表面标志物特征对 ADMSC 进行了表征。本研究使用了特定的成脂和成骨分化方案。通过实时 PCR 研究了天然 ADMSC 以及用不同浓度的维生素 D3（0、50、100、150 nM）和 VPA（0.5、1、1.5、2 mM）连续刺激 5 天后几种多能性、内胚层、胰腺祖细胞和胰腺内分泌谱系标志物的基因表达。此外，通过 ELISA 和免疫细胞化学法证实了生长抑素的表达。
在 ADMSC 中，检测到生长抑素 mRNA、维生素 D 受体（VDR）及其代谢酶 1α-羟化酶、24-羟化酶和 25-羟化酶的表达。用维生素 D3 刺激后，观察到维生素 D 受体（VDR）的核转位。有趣的是，维生素 D3 的存在降低了生长抑素基因的转录。相比之下，VPA 处理培养的 ADMSC 显示出对生长抑素基因表达的增强作用。在维生素 D3 和 VPA 的影响下，没有其他多能性、内胚层、胰腺祖细胞或胰腺内分泌谱系 mRNA 表达被调节。
人 ADMSC 携带 VDR。维生素 D 代谢酶 25-羟化酶对维生素 D3 的添加有反应。此外，我们的结果表明 ADMSC 中的生长抑素表达是组成型的，部分分泌的，并受维生素 D3 和 VPA 的调节。
