The Effect Of Hydrogen Sulfide In Autophagic Dysfunction Caused By Low Shear Stress In HUVECs

Background and Purpose:Some current research concluded that autophagy has a protective effect in atherosclerosis. In the development of atherosclerosis, the vascular endothelial cells autophagy is almost widespread. When autophagy is suppressed in vascular endothelial cells, vascular endothelial cells damaged thereby contributing to the development of As. In vascular bifurcation, the inside of blood vessels and vascular bending branches, As lesions and endothelial cells injury is the most easy to find. In these parts, hemodynamic characteristics have changed dramatically, resulting in abnormal shear stress. Although studies have shown that high shear stress or low shear stress can promote the formation of As. However, pathogenic conditions of high shear stress has far exceeded the normal physiological levels, many scholars prefer low shear stress is the more important factor. Studies have shown that low shear stress will suppress autophagy in vascular endothelial cells.H₂S（hydrogen sulfide, H₂S） is the third endogenous gaseous signaling molecule was discovered in recent years, it has a variety of biological effects, including anti-endothelial cell apoptosis, promote angiogenesis, anti-oxidation and anti-As. Studies have shown that hydrogen sulfide is capable of inducing a protective effect of autophagy through AMPK pathway, inhibition of cancerous colonic epithelial cell proliferation. Therefore, this study treated AMPK as astarting point, through the establishment of a low shear stress-induced autophagy dysfunction model in HUVECs to investigate the new mechanism that how H₂ S suppressed autophagic dysfunction in autophagy vascular endothelial cells caused by low shear stress.Method and Result:1, Ligaturing the left carotid artery and retain smaller thyroid artery in mice to established an animal experiment model. Protein expression is detected by immunofluorescence. In cells experiment we use HUVECs and treated it 1 hour by the normal shear stress（15dyne/cm²） and low shear stress（5dyne/cm²）, using western blot assay to examine CSE protein expression. The results showed that compared with the normal treatment group, protein expression of low shear stress group, were significantly lower.2, HUVECs was treated by normal shear stress（15dyne / cm²） and low shear stress（5dyne / cm²） 1 hour. The results show that t the protein expression of LC3 and Beclin 1 has significantly reduced and P62 was increased.3, HUVECs was treated with different concentrations of Na HS（0μmol / L, 25μmol / L,50μmol / L,100μmol / L）. The results from western blot assay showed that compared with the 0μmol / L group, H₂ S can antagonize autophagy caused by low shear stress. This antagonism in a concentration-dependent increase.4, The cells were randomly divided into three groups: normal shear stress15 dy / cm² group, low shear stress5 dy / cm² group,5mmol / L PPG + low shear stress group. Wherein the PPG pretreatment of cells 30 min, then using shear stress treatment 1h. Western blot assay results showed that the level of autophagy was reduced compared with5 dy / cm² group.5, The cells were randomly divided into four groups: normal shear stress15 dy / cm² group, low shear stress5 dy / cm² group,100μmol / L Na HS +5dy / cm² group,5mmol / L PPG +5dy / cm² group. Compared with the normal group, the level of p-AMPK / AMPK expression was increased and m TOR expression was decreased in5 dy / cm² group and PPG +5dy / cm² group.6, The cells were randomly divided into5 groups, normal shear stress15 dy / cm² group, low shear stress5 dy / cm² group,100μmol / L Na HS +5dy / cm² group, AMPK inhibitor Compound C +5dy / 2 group, Compound C +100μmol / L Na HS +5dy / cm² group. Comparing with5 dy / cm² group, the level of p-AMPK / AMPK and LC3 was increased and mTOR was decreased in the Na HS +5dy / cm² group. Expression of LC3 protein was decreased and m TOR expression increased in Compound C +5dy / cm² group. Comparing with Na HS+5dyne/cm² group, protein expression of p-AMPK/AMPK and LC3 was reduced and m TOR expression increased in Compound C + Na HS +5dy / cm² group.Conclusion:Low shear stress-induced autophagy dysfunction in HUVECs, and H₂ S can antagonize it. The mechanism of the inhibiting effect is involved with the activation of AMPK. Thereby reducing the intracellular levels of m TOR, and increased autophagy related proteins downstream.