[Shenqi Dihuang Decoction inhibits high-glucose induced ferroptosis of renal tubular epithelial cells via Nrf2/HO-1/GPX4 pathway].

This study aims to explore the effects of Shenqi Dihuang Decoction on high-glucose induced ferroptosis and the nuclear factor erythroid 2-related factor 2(Nrf2)/heme oxygenase-1(HO-1)/glutathione peroxidase 4(GPX4) axis in human renal tubular epithelial cells(HK-2) and to clarify the underlying mechanism. The cell injury model was established by exposing HK-2 to high glucose, and the Shenqi Dihuang Decoction-medicated serum was prepared. The optimal concentration and intervention time of Shenqi Dihuang Decoction were determined. HK-2 were divided into normal, high glucose, and low-, medium-, and high-dose Shenqi Dihuang Decoction groups. After interventions, the cell proliferation rate in each group was determined and the cell morphology and mitochondrial ultrastructure were observed. Then, the levels of intracellular reactive oxygen species(ROS), ferrous ion(Fe~(2+)), glutathione(GSH), and malondialdehyde(MDA) and the protein levels of Nrf2, HO-1, GPX4, and xCT were measured. The optimal concentration and intervention time of Shenqi Dihuang Decoction-medicated serum were determined to be 10% and 24 h, respectively. Compared with the high glucose group, high-dose Shenqi Dihuang Decoction promoted the proliferation of HK-2. The cells in the low-, medium-, and high-dose Shenqi Dihuang Decoction groups presented tight arrangement, an increased cell count, improved morphology from a spindle-fiber shape to a cobblestone shape, and improved morphology and structure of mitochondrial membrane and cristae, compared with those in the high glucose group. Meanwhile, all the doses of Shenqi Dihuang Decoction inhibited ROS elevation to mitigate the peroxidation damage, lowered the Fe~(2+) and MDA levels and elevated the GSH level to inhibit lipid peroxidation, and activated the antioxidant pathway to upregulate the protein levels of Nrf2, HO-1, xCT, and GPX4. In conclusion, Shenqi Dihuang Decoction-medicated serum can inhibit high-glucose induced ferroptosis of HK-2 in vitro, which involves the antioxidant effect and the activation of the Nrf2/HO-1/GPX4 pathway.

Astragaloside IV drug-loaded exosomes (AS-IV EXOs) derived from endothelial progenitor cells improve the viability and tube formation in high-glucose impaired human endothelial cells by promoting miR-214 expression.

INTRODUCTION: The high glucose changes caused by diabetes mellitus (DM) can damage the vascular system. Astragaloside IV (AS-IV) can improve diabetes and promote angiogenesis. Exosomes (EXOs) help to carry specific drugs into cells efficiently. However, whether AS-IV loaded EXOs (AS-IV EXOs) can improve damaged endothelial cells through miR-214 remains to be determined. MATERIAL AND METHODS: We prepared and identified AS-IV EXOs derived from endothelial progenitor cells (EPCs) and high glucose stimulated endothelial cell models to investigate whether AS-IV EXOs can improve damaged endothelial cells through miR-214. We used a transmission electron microscope (TEM) and DAPI staining to identify the morphology and characteristic expression of EPCs and EXOs, and then prepared AS-IV EXOs. Cell function tests were performed to detect the cloning, proliferation, and migration capabilities of cells. Western blot (WB) and real-time quantitative polymerase chain reaction (qRT-PCR) were used to assess the expression level of Tie-2, Ang-1, and PI3K/Akt-related protein. RESULTS: The DAPI staining results showed that inducing human umbilical vein endothelial cells (HUVECs) could effectively absorb AS-IV EXOs. The results of plate clone formation assay, CCK-8, cell adhesion, and transwell assay of HUVECs stimulated by high glucose showed that AS-IV EXOs had a damage relief effect. By the detection of WB and qRT-PCR, it was found that AS-IV EXOs promoted the expression of miR-214 and proteins related to blood vessel growth. After transfection of miR-214 to pre-treat HUVECs under high glucose stimulation, AS-IV EXOs promoted the tube formation of HUVECs by regulating the level of miR-214. CONCLUSIONS: By promoting the expression of miR-214, AS-IV EXOs significantly improved the activity and tubularization of HUVECs under high glucose stimulation.