Effects of glucagon-like peptide-1 analogue liraglutide on autophagy of human umbilical vein endothelial cells in high glucose conditions
10.11855/j.issn.0577-7402.2019.02.04
- Author:
Xiao-Xia LIU
1
Author Information
1. Department of Endocrinology, the Second Hospital of Shanxi Medical University
- Publication Type:Journal Article
- Keywords:
AMPK/mTOR;
Autophagy;
GLP-1 analogue;
High glucose;
Human umbilical vein endothelial cells;
Liraglutide
- From:
Medical Journal of Chinese People's Liberation Army
2019;44(2):113-119
- CountryChina
- Language:Chinese
-
Abstract:
Objective To explore the effect of glucagon-like peptide-1 (GLP-1) analogue liraglutide (LIRA) on autophagy of human umbilical vein endothelial cells (HUVECs) under high glucose condition, and its corresponding mechanism. Methods (1) The HUVECs were cultured in vitro and assigned to control group (group A) and high glucose group (group B). Group A and B were cultured respectively in 5.5mmol/L and 25mmol/L glucose medium for 12, 24 and 48 hours. The expressions of autophagy-related genes (beclin-1, LC3 and p62 mRNA) were determined by RT-PCR, and then the optimal culture time of high glucose condition on autophagy was selected. (2) The high glucose group at the optimal culture time point was treated with low, medium and high dose (10, 50, 100nmol/L) of LIRA (C1, C2, C3 groups). The same method was used to detect the indicators above, and the optimal intervention concentrations of LIRA were obtained. (3) In addition to the treatment of LIRA at optimal dose under high glucose condition (group C) and high glucose culture alone (group B), the AMPK inhibitor ComC (10μmol/L) was introduced (group D and group E). The fluorescence spots of green fluorescent protein (GFP) conjugated LC3 (GFP-LC3) were observed by laser confocal fluorescence microscopy to monitor the formation of autophagosome. The expressions of cell autophagy marker beclin-1, p62 and the ratio change of LC3-Ⅱ/LC3-, p-AMPK/AMPK and p-mTOR/mTOR were measured by Western blotting. Results (1) Compared with group A, the expressions of beclin-1 and LC3 mRNA in group B decreased, while of p62 mRNA increased at different culture time (P<0.05). The changes above were most significant at 24h, so the 24h was chosen as the optimal culture time. (2) Compared with group B, the expressions of beclin-1 and LC3 mRNA increased and of p62 mRNA decreased in C3 group (P<0.05), and the expressions of LC3 mRNA in C1, C2 and C3 groups all increased (P<0.05) in a dose-dependent pattern, so the optimal intervention concentration of LIRA was 100nmol/L. (3) The expressions of protein beclin-1, LC3-Ⅱ/LC3- and p-AMPK/AMPK increased, and of protein p62 and p-mTOR/mTOR decreased in group C compared with that in group B (P<0.05). The ratio of LC3-Ⅱ/LC3- and p-AMPK/AMPK was lower and of p-mTOR/mTOR was higher in group D than in group C (P<0.05). There was no significant difference in the expression level of protein beclin-1 and p62 between group C and D (P>0.05). Compared with group D, LC3-Ⅱ/LC3- ratio decreased and the expression of p62 protein increased in group E (P<0.05). The change trend of autophagy fluorescent spots GFP-LC3 was consistent with that of autophagy related protein. Conclusion GLP-1 analogue liraglutide may protect HUVECs from high glucose condition by activating autophagy, which was achieved through AMPK-mediated signaling pathway.
