Protective Effect Of Laminaria Polysaccharide On Vascular Endothelial Cells In Rats Induced By Psychological Stress

Social and psychological factors are the main reasons of inducing cardiovascular and cerebrovascular disease though damaging vascular endothelial cell （VEC）. Stressful situations, such as mental strains, anxiety,make dysfunction of the autonomic nervous system, sympathetic nerves activity,and adrenaline （Adr） releasing from adrenal medulla. After Adr broken down,the toxic metabolites can damage the morphological structure and the function of VEC. This study divides into five parts to explore the protective effect of laminaria polysaccharide on VEC.Part1 Exploration of an animal model of damaged VEC induced by psychological stressAim: To establish an animal model of damaged VEC induced by anger stress and evaluate the injury of VEC.Methods: To set up the animal model of endothelial cell injured by anger stress, adult male SD rats were randomly divided into control group and model group. The plasma levels of cortisol were detected by Radioimmunoassay, the plasma levels of Adr and vWF were detected by ELISA. The plasma NO levels were detected by nitrate reductase assay. The morphology of thoracic aorta endothelium was observed by HE stain and scanning electron microscopy（SEM）.Results: Compared with control group, the vertical, level and total scores increased significantly in model group. The plasma levels of cortisol, Adr and vWF in the model group were significantly higher than those of control group（P<0.05）. The plasma levels of NO was significantly lower than that of the control group （P<0.05）. And the morphology of the endothelial cell was significantly impaired.Conclusions: Anger stress causes the injury of VEC.Part 2 Protective effect of laminaria polysaccharide on morphological structure of damaged VEC in rats induced by psychological stressAim: To explore the protective effect of laminaria polysaccharide on morphological structure of damaged VEC in rats induced by psychological stress.Methods: Normal adult rats were randomly divided into control group,laminaria polysaccharide high dose group （LP-H）, laminaria polysaccharide low dose group （LP-L） and low molecular heparin group. The model of damaged VEC was build by psychological stress. After stress, all animals were conducted the open-field tests and took plasma to detect the plasma cortisol and Adr. Then aortic were isolated for observed SEM and transmission electron microscope（TEM）. And the ratios of VEC apoptosis was estimated by TUNEL stain.Results: Compared with control group, the vertical, level and total scores,the cortisol and Adr levels in plasma increased significantly in model group,LP-H, LP-L and low molecular heparin groups （P<0.05）. The surface of VEC appeared contraction, uneven and the intercellular space increase, but the surface of those in LP-H was improved in SEM. VEC appeared mitochondria cristae vague,vacuole in model group, but VEC in LP-H appeared mitochondria clear,integrity in TEM. And the apoptotic cells can be found in model group, LP-H,LP-L and low molecular heparin groups （P<0.05）. But the apoptotic ratio in model group was significantly increased compared with LP-H.Conclusions: Laminaria polysaccharide has protective effect on morphological structure of damaged VEC in rats induced by psychological stress.Part 3 Regulative effect of laminaria polysaccharide on vascontriction and vasdilation factors released by damaged VEC in rats induced by psychological stressAim: To explore the regulative effect of laminaria polysaccharide on vascontriction and vasdilation factors released by damaged VEC of rats induced by psychological stress.Methods: Normal adult rats are randomly divided into control group, LP-H,LP-L and low molecular heparin group. The model of damaged VEC of rat was build by psychological stress. After that, NO, endothelin （ET）, 6-Keto-PGF1a and thromboxane B2 （TXB2） in rat plasma were tested. Aortic rings were isolated to observe the tensions of the relaxation induced by Ach and the contraction induced by NE. After adding L-NAME A, the tensions of the vascular relaxation induced by nitroglycerin and the vascular contraction induced by NE were recorded respectively.Results: Compared with model group, LP-H markedly increased the plasma level of NO and 6-Keto-PGF1a（P<0.05）, decreased the plasma level of vWF and ET （P<0.05）. Compared with model group, Ach-induced relaxation significantly increased and NE-induced contraction significantly decreased in LP-H （P<0.05）. After adding in L-NAME the relaxation of model, LP-L, LP-H and heparin groups induced by nitroglycerin were rather lower than normal group （P<0.05）. The contraction of model, LP-L, LP-H and heparin groups induced by NE were rather higher remarkably than normal group （P<0.05）. But as to model group, the contraction decreased significantly in LP-L and LP-H groups （P<0.05）.Conclusions: Laminaria polysaccharide can correct the unbalance of vascular constricting and relaxing factors released by VEC of rats induced by psychological stress. Therefore, this indicated its protective effect on VEC.Part 4 Effect of laminaria polysaccharide on the plasma metabolic products of Adr in rat induced by psychological stressAim: To explore the effect of laminaria polysaccharide on the plasma metabolic products of endogenous Adr in rat induced by psychological stress.Methods: Normal adult rats are randomly divided into control group, LP-H,LP-L and low molecular heparin group. The model of damaged VEC of rat was build by psychological stress. After that, SSAO level in vascular tissue and the levels of MAO, SSAO, H₂O₂, Methylamine and formaldehyde in plasma were tested respectively.Results: There was no significant difference in plasma MAO level in all groups （P>0.05）. There was no significant difference in tissue SSAO level in control, model and low molecular heparin groups （P>0.05）. But the tissue SSAO levels in LP-H and LP-L were lower than those of model group （P<0.05）. There was no significant difference in plasma SSAO level in control, model, LP-L and low molecular heparin group （P>0.05）. But the plasma SSAO level in LP-H was higher than those of model group （P<0.05）. Compared with model group, the plasma level of H₂O₂, formaldehyde and methylamine decreased significantly in LP-H（P<0.05）.Conclusions: Laminaria polysaccharide protects VEC through decreasing the plasma metabolic products of endogenous Adr.Part 5 Protective effect of laminar polysaccharide on VEC damaged by the metabolic product of Adr in vitroAim: To explore the effect of laminar polysaccharide on VEC damaged by the metabolic products of Adr in vitro.Methods: The aortic rings were isolated from normal rats in vitro. The aortic rings were divided into control group, model group, LP-H and LP-L.H₂O₂, formaldehyde or methylamine were added to the model group respectively, while LP-H and LP-L were like the model group but adding high or low dose LP at the same time. Control group were added of same amout of K-H only. The tensions of vascular relaxation induced by Ach and the tensions of contraction induced by NE were recorded respectively.Results: After adding H₂O₂, the relaxations of LP-H, LP-L, control group were obvious more than those of model group （P<0.05）and there was no difference in the contractions of LP-H, LP-L, control group and model group.After adding formaldehyde, the relaxations of LP-H, LP-L and model group were obvious more than those of control group, but the relaxations of LP-H were obvious lower than those of model group （P<0.05）. And the contractions of the control group, LP-H and LP-L were obvious lower than those of model group （P<0.05）. After adding methylamine, the relaxations were obvious more than those of model group and the contractions were obvious lower than those of model group in LP-H and control group （P<0.05）.Conclusions: Laminar polysaccharide regulates vascular endothelium-dependent relaxation and contraction through protective effect on VEC damaged by the metabolic products of Adr.