Study Of The Interaction Between Gigantol,Syringic Acid And High-glucose-induced Human Lens Epithelial Cells

Objective:Studies of the interaction mechanisms between gigantol,syringic acid(SA)and human lens epithelial cells(HLECs)were to investigate the effects of dendrobium on diabetic cataract(DC)through the mechanical properties of living cell morphology,which was to uncover the anti-DC mechanism of dendrobium.Methods:1.Transmission electron microscope(TEM)was used to detect the ultrastructures in high-glucose-induced HLECs treated with gigantol and SA,to show the detail changes of nuclear,mitochondria and endoplasmic reticulum in HLECs.2.Atomic force microscopy(AFM)was used to investigate the changes of surface morphology and cell stiffness of high-glucose-induced HLECs treated with gigantol and SA in living cells level,and the changes of mechanical properties of HLECs with the surface roughness and cell stiffness were further calculated and analyzed by AFM images;3.Surface-enhanced Raman spectroscopy(SERS)was used to analyze the structure of C-C chain and inter-chain of cell membrane phospholipids in high-glucose-induced HLECs treated with gigantol and SA,and the relative intensity of Raman spectroscopy was used to calculate the changes of living cell membrane fluidity in different groups;4.Confocal laser scanning microscopy(CLSM)was used to observe the effects of gigantol and SA on the changes of cytoskeletal protein F-actin in high-glucose-induced HLECs through immunofluorescence assay.Results:1.The results of TEM was shown that the intact organelles in the cytoplasm,the uniform chromatin in nuclei,and the gathered mitochondria with clear and complete crista in the control group;the obvious pathological features that the chromatin deposition in the nucleus,and the obvious swelling and blurred phenomena in mitochondria,the obvious swelling phenomenon in endoplasmic reticulum in model group;the greatly reduced the injury changes of clumping of chromatin in nuclei,the swelling and degeneration in mitochondria,and the crista of mitochondria became relatively clear and complete in gigantol and SA group compared with model group.2.The height of HLECs was 3.5±0.35μm in control group;1.8±0.25μm in model group,compared with control group,P<0.01;3.1±0.15μm in gigantol group,compared with model group,P<0.01;2.8±0.25μm in SA group,compared with model group,P<0.01;and 3.8±0.43μm in gigantol+SA group,compared with model group,P<0.01.And the roughness Ra and Rq were 112±3.25nm,155±2.98nm in control group;1444±2.16nm,176±2.67nm in model group,114±3.45nm,135±5.12nm in gigantol group;132±4.12nm,161±3.91nm in SA group;and 104±4.21nm,121±2.76nm in gigantol+SA group.The stiffness of HLECs was 5.6±0.56KPa in control group,11.6±0.91KPa in model group,7.5±0.45KPa in gigantol group,8.9±0.45KPa in SA group,and 7.51±0.31KPa in gigantol+SA group.The roughness and stiffness was increased in model group,compared with control group,P<0.01,and decreased in gigantol group,SA group,and gigantol+SA group,compared with model group,P<0.01.This showed that giantol and SA could change the negative effects of morphological properties of HLECs in the presence of high glucose.3.Through Raman spectroscopy,the lateral interaction between chains Stran and the longitudinal order-parameters in chains Slat of HLECs in model group was significantly increased by 25%and 25.9%,compared with control group,P<0.01,and this indicated that the liquidity of HLECs was decreased;in gigantol group,Stran and Slat were decreased by 16%and 20%,compared with model group,P<0.01;in SA group,Stran and Slat were decreased by 2.9%and 3%;in gigantol+SA group,Stran and Slat were decreased by 5%and 20.8%,compared with model group,P<0.01 This results indicated that gigantol and SA could increase the liquidity of HLECs.4.From the imunofluorescence assay,in control group,the arrangement of the cytoskeletal protein F-actin was in order,and closed to the cell membrane;in model group,the arrangement of the skeletal protein F-actin was diffuse and messy in cell,and in the edge of cells,the ratio of F-actin was decreased;in gigantol group and SA group,the order of cytoskeletal protein F-actin in HLECs was showed the situation of recovery,which was indicated that gigantol and SA could be restored the order of cell cytoskeleton in HLECs.Conclusion:gigantol and SA could improve the pathological changes and the morphological changes in HLECs effectively.In this process,the C-C chain in lipid structure and cytoskeleton structure of cell membrane in HLECs were changed by gigantol and SA to increase the liguidity of HLECs to further decrease the surface roughness and cell stiffness of HLECs.Giangtol and SA played the therapeutic role role in DC through change the mechanical properties of cell morphology at real-time monitoring and living cells level.