Effect of insulin on the apoptosis of vascular endothelial cells induced by burn serum / 中华急诊医学杂志

Objective To investigate the effeets of insulin on the apoptosis of vascular endothelial cells cuinduced by burn$eruln in order to explore its possible mechamsm.Method Cultured human ECV304 cells were randomly divided into 33x)ups:control group,the ECV304 cells hured by 15%(V/V)rat normal,qertlm(t=6);bum semm group,the ECV3()4 cells simulated by 15%(V/V)self-made burn semm collected from rats with 30%TBSA full-thickness burns on the,back(n=6);and burn Serum+insulin group.the ECV304 cells cultared by insulin(10-7mol/L)and 15%(V/V) seIf-made rat bum serum(n=6).The transferase mediated nick end labeling(TUNEL) method was employed to measure the apoptosis of endothelial cells at 6 hours after stimulation.Meanwhile.immanohistochemical technique and Western blotting were used to determine the protein expressions of bcl-2 and eNOS.Data are expressed ills mean ±SEM.Statistical comparison was made using oneway analysis of vtriance.Significance was accepted at P<0.05.Results Compswith the control group,bum$erunl induced the apoptusis(18.5±3.1%)and down-regalated bcl·2(O.36±0.12)and p-eNOS(O.55±0.28)protein expressions of HUVECs(P<0.01).Burn 9AJ'unl+insulin significantly decreased the apoptosis(9.6 4-2.8%)and up-regulated bcl-2(0.944-0.25)and p-eNOS(0.89±0.16)protein expressions ofHU-VECs in comparison with the bum serllm group(P<0.01).eNOS showed no significant differences in three groups.Conclusions Insulin could markedly inhibit the apoptosis and up-regalate bcl-2 protein expression of HUVECs induced by bum serum,and its mec,harfism might involve the protein expression ofphosphorylated eNOS.

The screening and isolation of an effective anti-endotoxin monomer from Radix Paeoniae Rubra using affinity biosensor technology.

Lipopolysaccharide (LPS) is a known trigger in the pathogenesis of sepsis, lipid A being the toxic component. One of several adjuvant therapeutic approaches for severe sepsis is currently focusing on the neutralization of LPS. In order to obtain the components from traditional Chinese herbs that can neutralize the endotoxin, aqueous extractions were tested using affinity biosensor technology. From amongst 42 herbs, eight were found to possess lipid A-binding abilities. Radix Paeoniae Rubras had the highest lipid A-binding ability; therefore an aqueous extraction from this plant was investigated further. After preparation using standard methods, including silica gel chromatography and HPLC, we obtained 1, 2, 3, 4, 6-beta-d-pentagalloylglucose (PGG), with lipid A-binding ability. It was found that in vitro, PGG directly bound to lipid A, with a Kd of 32 microM, and that it neutralized the endotoxin both in the Limulus Amebocyte Lysate (LAL) assay and in a TNF-alpha release experiment, in a dose-dependent manner. In in vivo experiments, PGG was found to protect mice from a lethal challenge by LPS, and significantly decreased the plasma endotoxin level both in endotoxemic mice and rats, the reduction of the endotoxin level in rats being tightly associated with the TNF-alpha level. In conclusion, we demonstrate the effectiveness of affinity biosensor technology in discovering useful agents amongst traditional Chinese herbs and using this approach we found a new anti-endotoxin agent.

Inhibitory effects of intensive insulin on myocardium expression of tumor necrosis factor alpha in severely burnt rats / 解放军医学杂志

Objective To investigate the inhibitory effects of intensive insulin on myocardium expression of tumor necrosis factor alpha(TNF-?)in severely burnt rats and its correlative mechanism.Methods In eighteen SD rats the right jugular vein and the left ventricle were cathetetized,and were divided into three groups randomly:(1)the sham burn group,the rats were treated by sham burn;(2)the burn group,the rats were inflicted with 30%TBSA Ⅲ degree burn,and then given fluid resuscitation routinely;(3)the intensive insulin group,the rats received intensive insulin treatment after burn,and the liquid volume given was equal to the burn group.The level of plasma glucose contents,the left ventricular systolic pressure(LVSP)and left ventricular end-diastolic pressure(LVEDP)were recorded at 1,2,3,4,5,6h after burn,and animals were sacrificed at 6 hours after burn.The level of TNF-? in myocardium tissue was assayed by ELISA,and mRNA expression was determined by real-time PCR.Western blot was used to detect I?B? phosphorylation and degradation in left ventricle tissue.Meanwhile,burn serum-challenged cardiomyocytes were also used to detect the levels of insulin and LY294002,a specific insulin activation inhibitor,on the phosphorylation of I?B?.Results The animals showed stress hyperglycemia after burns,the function of heart was damaged(LVSP degraded and LVEDP increased),the level of TNF and the expression of mRNA in the myocardium increased apparently,I?B? phosphorylation and degradation increased in left ventricle tissue and burn serum-challenged cardiomyocytes.Given the intensive insulin treatment to keep the blood glucose level between 4.5 to 5.2 mmol/L,the function of heart was significantly improved,the level of TNF-? was apparently lowered,and the I?B? phosphorylation and degradation were significantly decreased.LY294002 could abolish the inhibitory effect of insulin,and it acted on the I?B? phosphorylation on burn serum-challenged cardiomyocytes model.Conclusion Intensive insulin may decrease the activation of NF-?B through diminishing the I?B? phosphorylation and degradation,which inhibit the transcription and expression of TNF-? in myocardial cell,and improve the heart function accordingly.