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蒲公英根提取物诱导 HEK293 细胞内钙离子增加。

Dandelion Root Extract Induces Intracellular Ca Increases in HEK293 Cells.

机构信息

Department of Biosciences, Biotechnologies and Biopharmaceutics, University of Bari, 70126 Bari, Italy.

Department of Sciences, University of Basilicata, 85100 Potenza, Italy.

出版信息

Int J Mol Sci. 2018 Apr 7;19(4):1112. doi: 10.3390/ijms19041112.

Abstract

Dandelion Weber ex F.H.Wigg.) has been used for centuries as an ethnomedical remedy. Nonetheless, the extensive use of different kinds of dandelion extracts and preparations is based on empirical findings. Some of the tissue-specific effects reported for diverse dandelion extracts may result from their action on intracellular signaling cascades. Therefore, the aim of this study was to evaluate the effects of an ethanolic dandelion root extract (DRE) on Ca signaling in human embryonic kidney (HEK) 293 cells. The cytotoxicity of increasing doses of crude DRE was determined by the Calcein viability assay. Fura-2 and the fluorescence resonance energy transfer (FRET)-based probe ERD1 were used to measure cytoplasmic and intraluminal endoplasmic reticulum (ER) Ca levels, respectively. Furthermore, a green fluorescent protein (GFP)-based probe was used to monitor phospholipase C (PLC) activation (pleckstrin homology [PH]-PLCδ-GFP). DRE (10-400 µg/mL) exposure, in the presence of external Ca, dose-dependently increased intracellular Ca levels. The DRE-induced Ca increase was significantly reduced in the absence of extracellular Ca. In addition, DRE caused a significant Ca release from the ER of intact cells and a concomitant translocation of PH-PLCδ-GFP. In conclusion, DRE directly activates both the release of Ca from internal stores and a significant Ca influx at the plasma membrane. The resulting high Ca levels within the cell seem to directly stimulate PLC activity.

摘要

蒲公英(Weber ex F.H.Wigg.)已被用作民族医学疗法数百年。尽管如此,不同种类的蒲公英提取物和制剂的广泛使用是基于经验发现的。一些报告的不同蒲公英提取物的组织特异性作用可能是由于它们对细胞内信号级联的作用。因此,本研究旨在评估乙醇蒲公英根提取物(DRE)对人胚肾(HEK)293 细胞钙信号的影响。通过 Calcein 活力测定法确定粗 DRE 增加剂量的细胞毒性。使用 Fura-2 和基于荧光共振能量转移(FRET)的探针 ERD1 分别测量细胞质和腔内质网(ER)内 Ca 水平。此外,使用绿色荧光蛋白(GFP)基探针监测磷脂酶 C(PLC)激活(pleckstrin 同源[PH]-PLCδ-GFP)。在存在外部 Ca 的情况下,DRE(10-400μg/mL)暴露剂量依赖性地增加细胞内 Ca 水平。在不存在细胞外 Ca 的情况下,DRE 诱导的 Ca 增加明显减少。此外,DRE 导致完整细胞中 ER 中 Ca 的显著释放和 PH-PLCδ-GFP 的共定位。总之,DRE 直接激活细胞内储存库中 Ca 的释放和质膜上的 Ca 内流。细胞内的高 Ca 水平似乎直接刺激 PLC 活性。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cba8/5979456/afe05b718ba6/ijms-19-01112-g001.jpg

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