| Background: Cardiovascular disease caused by atherosclerosis is a common complication and the main cause of death in patients with chronic renal failure. Patients with chronic renal failure displayed oxidative stress and "micro inflammatory state" in endothelial cells and the interaction between them was indicated as the pathology basis for endothelial injury.High concentration of urea was demonstrated to performing very important role in the ROS production, oxidative stress and disfunction of endothelial cells, but the molecular mechanism was not clear. The adaptor protein p66 Shc is a key protein in regulating oxidative stress, which induced oxidative stress and inflammation formation in endothelial cells by high glucose and ultraviolet ray. Unfortunately, the role of p66 Shc in urea-induced oxidative stress in endothelial cells has not been reported.Therefore, the investigaton the effect of p66 Shc on reactive oxygen species production and inflammation formation induced by urea in human umbilical vein endothelial cells may reveal the pathologic mechanism of cardiovascular disease in chronic renal failureObjective: To observe the production of ROS, changed expression pattern and level of p66 shc, NF-κB induced by urea in the HUVECs cells,to investigate mechanism and the effect of p66 Shc on reactive oxygen species production and inflammation formation induced by urea in human umbilical vein endothelial cells.Methods: Cultured human umbilical vein endothelial cells in vitro were randomly divided into normal group, control group and urea group.HUVECs cultured in vitro were incubated at different time points in urea(25 mmol/L) or mannitol(as the same osmotic control), Reactive oxygen species(ROS) were measured by ROS assay kit. Expression pattern of adaptor protein p66 Shc and NF-κB in the cells were observed by immunofluorescence assay. Protein expression levels of p66 Shc,p66Shc-Ser36, NF-κB/p65, p-p65 in HUVECs were detected by Western blot.Results: HUVECs used in this study were expressed FVIII widely and had good cell viability. Compared with the normal group and the mannitol control group, ROS were greatly enhanced after urea loading 1 h(P<0.05)and increased with time extention at 3 h(P<0.05). Immunofluorescence showed that high positive signaling of p-p66 Shc were detected in cytoplasm and nucleus after urea stimulued 3 h compared with control group and the mannitol control group. The positive expression of NF-κB/p65 in cytoplasm of HUVECs increased greatly. The expressionlevel of p66 Shc were increased in a time-dependent pattern and reached to peak at 24 h(P<0.05)by stimulation of 25 mmol/L urea on HUVECs,meanwhile enhancing the phosphorylation of p66Shc-Ser36(p<0.05);expression level of p-p65 were significantly increased at 3 h、6 h after urea treatment, reached to the peak at 12 h and decreaed at 24 h(P<0.05).Conclusion: p66 Shc may be involved in the pathogenesis of ROS overproduction induced by urea loading on HUVECs cells. Interaction of p66 Shc with ROS activated NF-κB pathway and resulting in inflammation,which may be play an important role in the cardiovascular diseases of the chronic renal failure.Background: Active or passive sleep deprivation is popular in modern life. The increasing of energy consumption caused by sleep deprivation is closely related to oxidative stress, and the latter is the main cause for inflammation induced by activation of tumor necrosis factor TNF-α. Pancreas is an important organ related to regulation enegy.However, the effect of oxidative stress on pancreatic morphology and function has not been reported by sleep deprivation. Detected the effect on pancreas by sleep deprivation maybe provide experimental evidence for chaging unhealthy life style and prevention pancreatic injury.Objective: To investigate the impact of sleep deprivation on morphology and function of pancreas in mice and the pathological significance of expression of tumor necrosis factor-a(TNF-a).Methods: C57 mice were randomly divided into four groups: control group, groups with sleep deprivation for 24 h, 48 h, 60 h, respectively. The mice subjected to sleep deprivation were observed for the general state and changes of body weight; serum amylase(serum amylase, AMS) level;Sectioned Pancreatic slices were observed by H&E staining and immunohistochemical staining to test the histological changes leaded from sleep deprivation and testing the expression of TNF-a in pancreatic tissue;Western Blot assay was practiced to detect the changing expression of TNF-a in the pancreas.Results: With the extension of time, mental state, the body weight of sleep deprived mice was significantly decreased compared with the control group. AMS levels in sleep deprived mice significantly increased in 24 h,reaching peak at 48 h and lasted till 60 h. H&E staining showed that mice deprived of sleep appeared turbid including cytoplast pyknosis of pancreatic islet cells, expansion of interstitial tissue; due to secretions of zymogen granules in acinar cells which induced the vacuolization of acinar cells. Immunohistochemistry and Western blot showed that sleep deprivation group had increased expression of TNF-a in pancreas with the extension of deprivation.Conclusion: Sleep deprivation leads to systemic failure in mice, and activates TNF-a-induced inflammation of the pancreas, leading to its morphological and functional damage. |
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