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Protective Effects of Oxymatrine on Vascular Endothelial Cells from High-Glucose-Induced Cytotoxicity by Inhibiting the Expression of A2B Receptor
- Source :
- Cellular Physiology and Biochemistry, Vol 45, Iss 2, Pp 558-571 (2018)
- Publication Year :
- 2017
-
Abstract
- Background/Aims: Diabetes mellitus (DM) has become an increasingly epidemic metabolic disease. Vascular endothelial cells play a key role in developing the cardiovascular complications of DM. The A2B receptor is expressed in vascular endothelial cells, and may help regulate the function of endothelial cells. The aim of this study was to investigate the protective effects of oxymatrine (OMT) on human umbilical vein endothelial cells (HUVECs) from high glucose-induced cytotoxicity. Methods: Homology modeling and molecular docking analysis were used to detect the binding sites between the adenosine A2B receptor and OMT. HUVECs were cultured with control (5.5 mM) or elevated glucose (22.2 mM) in the presence or absence of 3 µM OMT or A2B siRNA for 3 days. The MTS cell viability assay was used to measure the toxicity of high glucose on HUVECs and the protective effect of OMT or A2B siRNA. The expression of the adenosine A2B receptor and CCL5 in HUVECs was detected with real-time quantitative PCR (qPCR) and Western blotting methods in each group. Levels of IL-1β and TNF-α were measured using an enzyme-linked immunosorbent assay (ELISA) kit, and the concentration of NO was detected with the nitrate reductase method. Monocyte chemotactic activity in each group was detected using Transwell chambers. Furthermore, the phosphorylation of p38 and ERK1/2 in each group was observed through the Western blotting method. Results: Homology modeling and molecular docking analysis showed that OMT contains well-fitted binding sites to the A2B receptor. After chronic culture at high glucose, the rate of cell viability was significantly lower than that of the control group. After co-treatment with OMT or A2B siRNA, cell viability was significantly increased compared with the high-glucose group. The results from real-time quantitative RT-PCR (qRT-PCR) and Western blotting indicated that high glucose could increase the expression of A2B receptors in HUVECs, an effect that was inhibited by OMT. In addition, the results revealed that the expression of CCL5, IL-1β and TNF-α was increased in the high-glucose group, and that the NO produced by HUVECs decreased due to hyperglycemia; however, co-culture with OMT or A2B siRNA abolished these effects. Meanwhile, the chemotaxis activity of monocytes to HUVECs cultured in high-glucose medium was enhanced 2.59-fold compared to the control cells. However, the inflammatory reactions in HUVECs were completely relieved by co-treatment with OMT or A2B siRNA. Moreover, the phosphorylation of p38 and ERK1/2 in HUVECs in the high-glucose group was significantly higher than that of the control group; these effects were reversed after co-treatment with OMT or A2B siRNA. Conclusion: OMT may protect the HUVECs from high glucose-induced cytotoxicity through inhibitting the expression of A2B receptor and inflammatory factors as well as decreasing the phosphorylation of p38 and ERK1/2.
- Subjects :
- 0301 basic medicine
Physiology
Cell Survival
Endothelial cells
Interleukin-1beta
Gene Expression
Oxymatrine
Protective Agents
Receptor, Adenosine A2B
p38 Mitogen-Activated Protein Kinases
lcsh:Physiology
Umbilical vein
Monocytes
A2B receptor
lcsh:Biochemistry
03 medical and health sciences
chemistry.chemical_compound
Diabetes mellitus
Alkaloids
Human umbilical vein
medicine
Human Umbilical Vein Endothelial Cells
Humans
lcsh:QD415-436
Viability assay
Phosphorylation
Cytotoxicity
Receptor
Chemokine CCL5
Mitogen-Activated Protein Kinase 1
Mitogen-Activated Protein Kinase 3
lcsh:QP1-981
Tumor Necrosis Factor-alpha
Monocyte
Molecular biology
Protein Structure, Tertiary
Blot
Molecular Docking Simulation
030104 developmental biology
medicine.anatomical_structure
Glucose
chemistry
RNA Interference
Adenosine A2B receptor
Quinolizines
Signal Transduction
Subjects
Details
- ISSN :
- 14219778
- Volume :
- 45
- Issue :
- 2
- Database :
- OpenAIRE
- Journal :
- Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology
- Accession number :
- edsair.doi.dedup.....616e51d5108782d6b000ce005d619989