Liver protection by Sini decoction in hemorrhagic shock and its mechanism relating to oxygen free radical and nitric oxide

AIM:The work was designed to explore protective effects of a traditional Chinese medicine-sini decoction (SD) on liver in hemorrhagic shock and its mechanism relating to oxygen free radical and nitric oxide.METHODS:Anesthetized Wistar rats were subjected to a hemorrhagic shock protocol for 60 min followed by intravenous injection with normal sodium chloride solution or SD solution. Superoxide dismutase (SOD), malondialdehyde (MDA) and nitric oxide (NO) in liver were examined. The inducible nitric oxide synthase (iNOS) was determined immunohistochemically. RT-polymerase chain reaction (RT-PCR)was used to assay the mRNA, which were corresponding to eNOS (endothelial nitric oxide synthase) and iNOS.RESULTS:The activity of SOD decreased, while the concentration of MDA increased in liver during hemorrhagic shock. SD enhanced SOD activity and inhibited a increase in MDA level in liver (P＜0.01). The NO concentrations in liver in SD group increased at three hours after resuscitation (P<0.01). In addition, it was found that the expression of iNOS was upregulated in sodium chloride-treated group, while SD upregulated the expression of eNOS.CONCLUSION:SD reduces the liver injury caused by oxygen free radicals during hemorrhagic shock. The increasing NO concentration by SD is through upregulation of endothelial NOS expression.     

Differential expression of Ang-1 and Ang-2 proteins contributes to biphasic changes of vascular reactivity after hemorrhagic shock in rats

AIM: To observe the protein expression of angiopoietin-1 (Ang-1) and angiopoietin-2 (Ang-2) after hemorrhagic shock, and to explore the role of this difference in the development of the biphasic change of vascular reactivity after hemorrhagic shock in rats. METHODS: The vascular reactivity of the first class arborization of superior mesenteric artery (SMA) and protein expression of Ang-1 and Ang-2 after hemorrhagic shock were measured via the isolated organ perfusion system and Western blotting technique. The relationship between the protein expression of Ang-1/Ang-2 and the vascular reactivity after hemorrhagic shock was observed. By treatment with Ang-1 and Ang-2 or inhibition of Ang-1 and Ang-2, the effects of angiopoietins on the vascular reactivity of SMA in the early stage (hyperreactivity) and late period (hyporeactivity) of hypoxia were observed to analyze the role of the differential expression of Ang-1 and Ang-2 protein in the biphasic change of vascular reactivity after hemorrhagic shock. RESULTS: (1) The vascular reactivity of SMA showed a biphasic change following hemorrhagic shock, which was increased in the early stage of shock, and decreased in prolonged stage. The E_max of norepinephrin in the shock 10 min group was the highest (129.3% of normal control, P<0.01). The protein expression of Ang-1 in SMA was also increased in the early stage of shock, decreased at prolonged stage of shock, and topped at 10 min shock group (1.85 folds of normal control, P<0.01). The protein expression of Ang-2 in SMA didn't change obviously in the early stage of shock, yet it was increased at prolonged stage of shock, and topped at 4 h shock group (2.90 folds of normal control, P<0.01). (2) In the hyperreactivity stage (hypoxia for 30 min), the extrinsic source of Ang-1 further increased the vascular reactivity, while the extrinsic source of Ang-2 decreased the vascular reactivity (P<0.01). In the hyporeactivity stage (hypoxia for 4 h), Ang-1 improved the vascular reactivity (P<0.01), while Ang-2 further decreased the vascular reactivity (P<0.01). CONCLUSION: The protein expression of Ang-1 and Ang-2 is different after hemorrhagic shock, and the differential expression of Ang-1 and Ang-2 contributes to the biphasic changes of vascular reactivity after hemorrhagic shock in rats.     

Effect of tea polyphenols on the contractile function of the papillary muscles of right ventricle in diabetic rats

AIM: To investigate the effect of tea polyphenols (TP) on the contractile function of the papillary muscles of right ventricle in the sixth and eighth week diabetic rats and the control rats. METHODS: Rats were induced to diabetic by an intravenous injection of alloxan. The papillary muscles of right ventricle in the sixth and eighth week diabetic rats were segregated and superfused with oxygenated Tyrode's solution. The contractile function and functional change under the electric stimulation state was recorded and compared between diabetic group and control group. RESULTS: 1/2 diastole interval elongation (P<0.05) and the depression of +dT/dtmax (P<0.01) and -dT/dtmax (P<0.05) of the papillary muscles of right ventricle were showed in the 6th week diabetic rats, while further damages, such as tension addition (P<0.01), 1/2 diastole interval elongation (P<0.01) and the depression of +dT/dtmax (P<0.01) and -dT/dtmax (P<0.01) in 8th week diabetic rats were also observed. TP at certain concentrations (15-120 mg/L) did not produce any effect on the 8th week diabetic rat's papillary muscle, but the positive muscle strength has been showed in the control rats and the sixth week diabetic rats. CONCLUSIONS: TP produces the positive muscle strength in the early diabetic cardiomyopathy, but has no beneficial effect on serious impaired diabetic myocardium.     

Pinacidil pretreatment increases vascular reactivity and calcium sensitivity after hemorrhagic shock

AIM: To observe the protective effects of protein kinase Cα(PKCα) and protein kinase Cε(PKCε) activated by pinacidil pretreatment on vascular reactivity and calcium sensitivity after hemorrhagic shock in rats. METHODS: The changes of the pressor effect(the change of mean arterial pressure) and vasoconstriction response(the changes of diameter) of superior mesenteric artery(SMA) to norepinephrine(NE) were observed. The vascular reactivity and calcium sensitivity of the first class arborization of SMA induced by pinacidil pretreatment with different volume and at different time points before shock were determined. The effects of PKCα and PKCε antagonists on the protection of pinacidil pretreatment, and the effects of pinacidil pretreatment on the translocation of PKCα and PKCε were also measured. RESULTS: (1) The pressor effect and vasoconstriction response of SMA to NE, and the vascular reactivity and calcium sensitivity of the first class arborization of SMA in 2 h shock group were significantly decreased as compared to those in normal controls(P<0.01). Pinacidil(25 μg/kg) pretreated at 30 min before shock attenuated the above changes.(2) The inhibitors of PKCα and PKCε suppressed the protective effects of pinacidil pretreatment(25 μg/kg pinacidil pretreated at 30 min before shock) on the vascular reactivity and calcium sensitivity. The E_max of NE was decreased by 42.9% and 62.9%, respectively(P<0.01). The E_max of Ca²⁺ was decreased by 31.1% and 56.1%, respectively(P<0.01). Pinacidil(25 μg/kg) pretreated at 30 min before shock increased the protein expression of PKCα and PKCε on the membrane, and decreased the protein expression in the cytoplasm as compared to those in 2 h shock group(P<0.01). CONCLUSION: Pinacidil pretreatment activates PKCα and PKCε, and induces the increasing effects of vascular reactivity and calcium sensitivity after hemorrhagic shock in rats.     

The damages of adrenal gland in rabbits with neisseria memingococcal septic shock

AIM:To investigate the damages of adrenal gland in rabbits infected with neisseria meningococci(MC).METHODS:18 rabbits were injected with MC (1×10¹¹ organism/kg, iv, model group, n=10) or normal saline (control group, n=8).TNF-α was detected in baseline (before challenged), 60 min and 120 min after injection. 120 min after injection, TNF-α immunochemical localization of adrenal gland was detected with ABC methods by electron microscopy. RESULTS:In model group, TNF-α positive cells were located in medulla and TNF-α released into simus of medulla. The medulla damages were appeared but gland cells of cortex was normal. No damages were showed in control group. TNF-α value of plasm was elevated at 60 min and 120 min after injection compared with control group. CONCLUSION:TNF-α was released into medulla and resulted in medulla damages after MC challenging. Adrenal cortex didnt damaged in early phase of MC septic shock.     

Effect of Shenmai injection on cardiomyocyte apoptosis after acute anoxia-reoxygenation

AIM: To study the effects of Shenmai injection on cardiomyocytes apoptosis after acute anoxia-reoxygenation (A/R) and the possible mechanism. METHODS: In this experiment, cultured cardiomyocytes isolated from neonatal rat were used. Model of myocardial anoxia-reoxygenation injury was produced by depriving oxygen for 5 min and then restoring oxygen for 15 min. The apoptotic cells was detected by flow cytometry to detect labbled Annexin V-FITC/PI. The intracellular calcium level was observed by laser scanning confocal microscopy markered Fluo-3/AM. RESULTS: In anoxia-reoxygenation group, the percentage of apoptotic cells and fluorescent intensity of intracellular calcium were both prominently higher than those in control group (P<0.01). The apoptotic rate in Shenmai injection group was notably less than that in A/R group and the intracellular calcium overload was also less obvious in Shenmai injection group than that in A/R group (P<0.01). CONCLUSION: Shenmai injection has notable effects on attenuating apoptotic rate after acute anoxia-reoxygenation in cardiomyocytes, which may be partly due to its alleviating intracellular calcium overload.     

Change of Gq protein-phosphatidyl inositol signaling in brain of rats with acute respiratory distress syndrome

AIM: To study the change of Gq protein-phosphatidyl inositol signaling in the brain of rats with acute respiratory distress syndrome (ARDS). METHODS: Forty male Wistar rats were randomly divided into oleic acid groups (OA groups of 30 min, 60 min, 90 min and 120 min exposure) and control group. The ARDS model in rats was reproduced by intravenous injection of oleic acid (0.2 mL/kg in 2 min). The mean arterial blood pressure (MABP), blood gas indexes, malondialdehyde (MDA), lactate dehydrogenase (LDH) and creatine kinase (CK) in plasma and brain tissues were measured. Gαq/11 protein and phospholipase C in brain tissues were detected by Western blotting. RESULTS: Compared to control group, MABP and PaO2 in all OA groups obviously decreased (P<0.05). MDA concentration and LDH activity in plasma and brain tissues of OA rats at time points of 90 min and 120 min were significantly higher than those in the controls (P<0.05). The CK activity in plasma of all OA groups increased (P<0.05). The CK activity in the brain tissues peaked at 60 min after OA exposure (P<0.05) and then decreased at 90 min and 120 min after treated with OA (P<0.05). Compared to control group, the concentration of Gαq/11 protein in the brain tissues of OA 60 min, 90 min and 120 min groups, and PLC concentration in the brain tissues of all OA groups obviously increased (P<0.05). A negative correlation between the change of Gαq/11 protein in the brain tissues and PaO2 (r=-0.579, P<0.05), a positive correlation between the change of Gαq/11 protein and MDA in the brain tissues (r=0.538, P<0.05), and a positive correlation between the change of Gαq/11 protein and LDH in the brain tissues (r=0.624, P<0.05) were observed. CONCLUSION: Up-regulation of Gq protein-phosphatidyl inositol signaling in the brain may play a role in the brain injury during ARDS.     

Astragalus injection inhibits the expression of JNK3 mRNA after hypoxia/hypoglycemia and reoxygenation in hippocampal neurons of rats

AIM: To investigate the effect of astragalus injection on the expression of c-jun N terminal kinase (JNK3) mRNA interrelated with apoptosis after hypoxia/hypoglycemia and reoxygenation in hippocampal neurons of rats. METHODS: The hippocampal neurons cultured for eight days were divided into 4 groups: normal control group, original astragalus injection group, hypoxia/hypoglycemia and reoxygenation group, astragalus injection group. Hypoxia/ hypoglycemia and reoxygenation group, astragalus injection group and original astragalus injection group were treated with hypoglycemia and reoxygenation after deprived of oxygen and glucose for 30 min. Methods of in situ hybridization and RT-PCR were used respectively to measure the expression of JNK3 mRNA after hypoxia/hypoglycemia and reoxygenation 0 h, 0.5 h, 2 h, 6 h, 24 h, 72 h and 120 h. RESULTS: In normal control group the volume of hippocampal neuronal nucleolus was accretion, cellular tuber was distinct and cytokinesis was dyed by yellow a lot. In hypoxia/hypoglycemia and reoxygenation group the hippocampal neuronal nucleolus was crimple, cellular tuber was shrinked, large number of cytokinesis was dyed by yellow and yellow granule was observed. Compared with normal control group, the numbers of JNK3 mRNA positive neuronal cells at each time point increased obviously in the hypoxia/hypoglycemia and reoxygenation group (P<0.05). The change of neuronal configuration and the numbers of JNK3 mRNA positive neuronal cells in original astragalus injection group accorded with hypoxia/ hypoglycemia and reoxygenation group (P>0.05). In astragalus injection group the hippocampal neuronal nucleolus was crimple slightly and segmental cytokinesis was dyed by yellow.Compared to hypoxia/hypoglycemia and reoxygenation group, the numbers of JNK3 mRNA positive neuronal cells at each time point were less obviously in the astragalus injection group besides 120 h (P<0.05). Compared to normal control group, the mean optic density of expression of JNK3 mRNA in hippocampal neurons of rats at each time point increased obviously in hypoxia/ hypoglycemia and reoxygenation group (P<0.05). Compared to hypoxia/hypoglycemia and reoxygenation group, the mean optic density of JNK3 mRNA expression at each time point in original astragalus injection group had no obvious change (P>0.05), however the mean optic density of JNK3 mRNA expression in hippocampal neurons of rats at each time point decreased obviously in the astragalus injection group besides 120 h (P<0.05). CONCLUSION: Astragalus injection inhibits the expression of JNK3 mRNA after hypoxia/hypoglycemia and reoxygenation, accordingly inhibits hippocampal neuronal apoptosis.     

Effects of ligustrazine ferulate on myocardial apoptosis and apoptosis-related protein expression in rats with myocardial ischemia-reperfusion injury

AIM:To observe the effects of ligustrazine ferulate on the apoptosis of myocardial cells in rats with myocardial ischemia-reperfusion injury, and to explore its possible mechanism. METHODS:Sixty male SD rats were randomly divided into five groups: sham-operation group, ischemia-reperfusion group, ligustrazine (4 mg/kg) group, low-dose (4 mg/kg) ligustrazine ferulate group and high-dose (8 mg/kg) ligustrazine ferulate group. The rat myocardial ischemia-reperfusion model was established by 30 min of myocardial ischemia followed by 120 min of reperfusion. Drugs were administered to the rats by jugular vein injection 10 min before reperfusion. After the reperfusion was finished, the biochemical indicators in serum and the histological indexes in myocardium were detected. RESULTS: Compared with ischemia-reperfusion group, ligustrazine ferulate lowered the serum levels of creatine kinase MB form, lactate dehydrogenase, cardiac troponin I and malondialdehyde, elevated the activity of total superoxide dismutase in serum and the expression of Bcl-2 protein in myocardium, decreased the expression of Bax protein and myocardial apoptotic index, and increased the Bcl-2/Bax ratio (all P<0.01). All the indicators in ligustrazine ferulate groups were dose-dependently superior to those in ligustrazine group (P<0.05 or P<0.01). CONCLUSION: Ligustrazine ferulate protects rats against myocardial ischemia-reperfusion injury. Its anti-apoptotic effect may be related to up-regulation of Bcl-2 and down-regulation of Bax.     

The signal transducers involved in the regulation of calcium sensitivity of vascular smooth muscle in hemorrhagic shock

AIM: To observe the regulatory effects of Rho-kinase, PKC and PKG on calcium sensitivity of vascular smooth muscle in hemorrhagic shock in rats. METHODS: The superior mesenteric artery (SMA) from hemorrhagic shock model of rat was adopted to assay the calcium sensitivity via observing the contraction initiated by Ca2+ under depolarizing conditions (120 mmol/L K+) with isolated organ perfusion system. Rho-kinase agonist Ang-Ⅱ and inhibitor fasudil, PKC agonist PMA and inhibitor staurosporine, PKG agonist 8Br-cGMP and inhibitor KT-5823 were used as tool agents to study the regulatory effect of Rho-kinase, PKC and PKG on the calcium sensitivity of SMA following shock. RESULTS: Ang-Ⅱ, PMA and KT-5823 improved the calcium sensitivity of SMA and made the cumulative dose-response curve of SMA to Ca2+ shift to the left, their Emax of Ca2+ (at 3×10-2 mol/L) was 0.630 g/mg, 0.595 g/mg and 0.624 g/mg, respectively, which were all higher than that in shock control (0.377 g/mg) (P<0.05, P<0.01). Fasudil, staurosporine and 8Br-cGMP delimitated the calcium sensitivity of SMA and made the cumulative dose-response curve of Ca2+ shift to the right, their Emax at 3×10-2 mol/L of Ca2+ was 0.242 g/mg, 0.230 g/mg and 0.256 g/mg, respectively, which were all lower than that in shock control (0.377 g/mg) (P<0.05, P<0.01). CONCLUSION: Rho-kinase, PKC, PKG play important roles in the regulation of calcium sensitivity of vascular smooth muscle in hemorrhagic shock.     

Effects of atorvastatin on nitric oxide,endothelin-1 and myocardial function in a rabbit model of acute myocardial infarction and reperfusion

AIM: To evaluate the effects of atorvastatin on nitric oxide(NO), endothelin-1(ET-1)and myocardial no-reflow in a rabbit model of acute myocardial infarction and reperfusion(AMI/R). METHODS: Twenty-four rabbits were randomized into 3 groups: 8 in AMI/R group, 8 in atorvastatin-treated group(5 mg·kg-1·d-1)and 8 in sham-operated group. Animals in the former two groups were subjected to 60 min of coronary occlusion followed by 120 min of reperfusion. Data on haemodynamics were collected. NO in blood sample, and in normal, and in infarcted reflow and no-reflow myocardium were evaluated respectively by nitrate reductase method. The levels of ET-1 in blood sample, and in normal, infarcted reflow and no-reflow myocardium were determined by radioimmunoassay. RESULTS: (1)Compared to the baselines, the heart rate(HR), systolic blood pressure(SBP), diastolic blood pressure(DBP), left ventricular systolic pressure(LVSP), maximal rate of increase and decline in left ventricular pressure(±dp/dtmax)and cardiac output(CO)in AMI/R and atorvastatin-treated groups were significantly declined, whereas left ventricular end-diastolic pressure(LVEDP)was increased after 60 min of coronary occlusion and 120 min of reperfusion(P<0.05 or P<0.01). However, in atorvastatin-treated group, LVSP, LVEDP, ±dp/dtmax and CO at the time point of 120 min of reperfusion recovered more significantly than those at the time point of 60 min of coronary occlusion(P<0.01), which was more significant than those in AMI/R group(P<0.05 or P<0.01). Compared to AMI/R group, the SBP and DBP were significantly heigher in atorvastatin-treated group(P<0.01).(2)In atorvastatin-treated group, the levels of ET-1 in blood sample were significantly lower than those in AMI/R group(P<0.01), and the levels of NO were significantly higher(P<0.01). Moreover, the levels of NO or ET-1 in infarcted reflow myocardium were significantly lower than that in AMI/R group(P<0.05 or P<0.01).(3)Atorvastatin could ameliorate myocardial function. CONCLUSION: Atorvastatin is effective in increasing NO and reducing ET-1 in blood plasma and local myocardium, and in protection of endothelial cells. Atorvastatin also has a beneficial effect on improving left ventricular function during acute myocardial infarction and reperfusion in rabbits.     

Mechanisms of hepatocytic mitochondria damage following septic shock

AIM: To study mechanism of hepatocytic mitochondria damage following septic shock. METHODS: 30 SD rats were randomly divided into three groups: sham operation group, 12 h cecal ligation and puncture (CLP) group and 16 h CLP group. The model of septic shock was made by cecal ligation and puncture. The liver mitochondria respiratory control rate (RCR), phosphate/oxygen (P/O) and ATPase activities were assayed. RESULTS: In 12 h CLP group mean artery pressure (MAP) [(9.54±1.26)kPa] was significantly lower than sham operation group [(14.58±1.32)kPa,P<0.05]. However, mortality was obviously higher than sham operation group (P<0.05), the liver mitochondria respiratory control rate (1.27±0.25), phosphate/oxygen (1.67±0.34) and Na⁺-K⁺-ATPase (40.80±3.45), Ca²⁺-ATPase (58.00±2.43), Mg²⁺-ATPase (78.30±4.16), Ca²⁺-Mg²⁺-ATPase(2.70±2.25) activities decreased strikingly. The difference between 12 h CLP group and sham operation group was significant (P<0.05), 16 h CLP groups was more lower than 12 h CLP group. As RCR, P/O and ATPase activities were significantly reduced, mortality significantly increased. Futhermore, obvious positive correlation was showed between them (r=0.892,P<0.01;r=0.834,P<0.01). CONCLUSION: Liver mitochondria function of ingestion-oxygen and phosphorus-acidification are decreased and membrane fluxion is weaken. Energy metabolism is blocked and Ca²⁺-Mg²⁺ shows imbalanced. All of them cause hepatocytic mitochondria injury following septic shock.     

Recovery of endothelial dysfunction with tolerogenic dendritic cell loaded with heat shock protein 60 in apolipoprotein E-null mice

AIM: To examine whether tolerogenic dendritic cells (DC) loaded with heat shock protein 60 (HSP60) could restore endothelial function in hypercholesterolemic apolipoprotein E (apoE)-null mice.METHODS: Bone marrow derived DC of the mice was loaded with HSP60 and co-cultured with rapamycin to generate tolerogenic DC.The tolerogenic DC, DC loaded only with HSP60 (DChsp) and saline were injected into the apoE-null mice at 6 weeks of age for two times at a one-week interval.C57BL/6 mice at the same age were taken as normal control two weeks after the last injection.Aorta was harvested for ex vivo vascular ring tension test.Immune parameters were also analyzed in vitro and in vivo.RESULTS: Compared with the non loaded DC, HSP60 pulsed DC expressed higher levels of CD86, and stimulated T lymphocytes to proliferation significantly, while the tolerogenic DC expressed lower levels of CD86, and inhibited T lymphocytes to proliferation.After immunization with different injection, Ach-induced relaxation was reduced significantly in DChsp group compared with saline group (P<0.01).Treatment of mice with tolerogenic DC restored endothelium-dependent dilation in a dose-dependent manner (P<0.01).The improvement in endothelial function was associated with a reduction in T cell response to HSP60.CONCLUSION: Our results indicate a rapid improvement in endothelial function with HSP60 tolerogenic DC immunization, and suggest that this immune therapy has significant vasculoprotective effects.     

Protective effects of diltiazem on liver,pancreas and small intestine in hemorrhagic-shock canine

AIM: To investigate the protective effects of diltiazem (Dil) on liver, pancreas and small intestines in hemorrhagic-shock canines and its mechanism. METHODS: The canines were bled to a mean arterial pressure (MAP) of 5.33-6.67 kPa for 30 min to produce the model of shock. During the shock, the dogs received water-soluble calcium channel blocker Dil or normal saline. The MAP was kept at the level for 90 min, then the total blood which was bled previously was reperfused. They were observed for 240 min. RESULTS: Dil could significantly elevated MAP of the hemorrhagic-shock canines (P<0.01) and the activity of superoxide dismmutase (SOD) of pancreas tissue (P<0.01), also it could decrease content of malondialdehyde (MDA) of the liver, pancreas and small intestine tissues (P<0.01). Electron microscope data indicated that Dil-treated dogs have a normal ultrastructure in the liver, pancreas and small intestine tissues. CONCLUSION: Dil can protect the structure and function of the liver, pancreas and small intestine in hemorrhagic-shock canines.     

Effect of Astragalus injection on expression of calmodulin after hypoxia/hypoglycemia and reoxygenation in rat hippocampal neurons

AIM: To investigate the effect of Astragalus injection on the expression of calmodulin(CaM) after hypoxia/ hypoglycemia and reoxygenation in rat hippocampal neurons.METHODS: The hippocampal neurons were cultured for 8 days and divided into 4 groups: normal control group (normal control), hypoxia/hypoglycemia and reoxygenation group (model), Astragalus injection solution group (solution control) and Astragalus injection group ( Astragalus ).The cells in all groups were treated with reoxygenation and normal medium after deprived of oxygen and glucose for 30 min except normal control group.The method of immunohistochemistry was used to measure the number of caspase-3 positive neurons.The expression of CaM at mRNA and protein levels was measured at time points of 0 h, 0.5 h, 2 h, 6 h, 24 h, 48 h, 72 h and 120 h after hypoxia/hypoglycemia and reoxygenation by RT-PCR and Western blotting, respectively.RESULTS: No difference of the parameters at all time points between model group and solution control group was found.Compared with normal control group, the numbers and the percentages of caspase-3 positive cells at all time points obviously increased in model group except at 0 h and 0.5 h (P<0.05).Compared with model group, the numbers and the percentages of caspase-3 positive cells were decreased in Astragalus injection group except at 0 h and 0.5 h (P<0.05).Compared with normal control group, the protein expression of CaM in rat hippocampal neurons at all time points obviously increased in model group (P<0.05).However, the protein expression of CaM in rat hippocampal neurons at all time points obviously decreased in Astragalus injection group as compared with model group (P<0.05).Compared with normal control group, the mRNA expression of CaM in rat hippocampal neurons at all time points obviously decreased in model group (P<0.05).The mRNA expression of CaM in rat hippocampal neurons at all time points obviously increased in Astragalus injection group as compared with model group (P<0.05).CONCLUSION: Astragalus injection inhibits the protein expression of CaM, the calcium overload and the expression of caspase-3 after hypoxia/hypoglycemia and reoxygenation, thus inhibiting hippocampal neuronal apoptosis.     

Effect of L-arginine on arrhythmia induced by ischemia/reperfusion of right coronary artery in rabbits

AIM: To explore the effect of L-arginine (L-Arg) on the arrhythmia induced by ischemia/reperfusion (IR) of the right coronary artery in rabbits. METHODS: 48 healthy adult rabbits were divided into 4 groups (n=12 of each) randomly: IRa group (120 min reperfusion after 30 min ischemia), IRb group (120 min reperfusion after 120 min ischemia), IRa+L-Arg group and IRb+L-Arg group. I/R model was established by occluding and loosening the root of the right coronary artery in rabbits. The changes of ECG and arrhythmia were recorded and graded. RESULTS: ① The longer time of IR was, the higher the score of the arrhythmia was found. The incidence of atrial-ventricular block (AVB), sinus-atrial block (SAB), even sinus arrest were detected and aggravated gradually. ② The incidence of AVB was decreased and from Ⅲ°→Ⅱ°→Ⅰ° markedly, some of sinus and atrial arrhythmia were transformed into sinus rhythm gradually, and all of the arrhythmia scores in IRa group were decreased significantly as compared with the same time phases of IRb group (P<0.01). ③ All of the arrhythmia scores in IRa+L-Arg and IRb+L-Arg groups were decreased dramatically as compared with that in IRa and IRb groups at the same time phases (P<0.01). ④ All of the arrhythmia scores in IRa+L-Arg group were lower compared with those in IRb+L-Arg group (P<0.01). CONCLUSION: Supplying appropriate L-arginine to the tissue is beneficial for inhibiting arrhythmia during ischemia and reperfusion, and the longer the time of ischemia is, the weaker the effect of L-arginine on the arrhythmia presented during the period of reperfusion.     

Effects of tirofiban on no-reflow and activation of NF-κB after myocardial ischemia/reperfusion in rats

AIM: To investigate the effects of platelet glycoprotein Ⅱb/Ⅲa receptor inhibitor tirofiban on myocardial no-reflow and activation of NF-κB after acute ischemia/reperfusion in rats. METHODS: Male Wistar rats were randomized into sham operation group, control group and tirofiban treatment group. Control group and tirofiban group were subjected ischemia for 90 min by ligation of coronary artery after thoracotomy and subsequently reperfusion for 120 min to establish acute myocardial ischemia/reperfusion no-reflow models. Thioflavine S, Evans blue and triphenyltetra zolium chloride (TTC) staining were performed to evaluate the area of no-reflow (ANR), infracted area (IA) and risk area (RA) of the heart. Immunohistochemistry was used for semi-quantitative analysis of the expression of nuclear factor-κB p65 (NF-κB p65) protein in myocytes and arteriole. Activity of myeloperoxidase (MPO) and content of malondialdehyde (MDA) in risk area of the heart were detected by ultraviolet spectrophotometer. RESULTS: After 120 min for reperfusion, compared to sham group, the statistical differences of higher positive expression of NF-κB p65 in myocytes and arteriole, activity of MPO and content of MDA both in control and tirofiban group were observed. Compared to control group, lower positive expression of NF-κB p65 in myocyte and arteriole, activity of MPO and content of MDA in tirofiban group were found (P<0.05, P<0.01). A markedly reduced ANR and IA were observed in tirofiban group than those in control group (34.36%±6.04% vs 52.09%±6.89%, P<0.01; 80.41%±8.48% vs 90.13%±5.72%, P<0.05). CONCLUSION: After myocardial ischemia/reperfusion for 120 min, no-reflow phenomenon can be observed in rats. Tirofiban reduces the areas of anatomic no-reflow and infarction, inhibits the activation of NF-κB in myocyte and arteriole, and decreases the infiltration of neutrophils and release of oxygen free radicals.     

Effect of ischemic preconditioning on vascular reactivity and calcium sensitivity in hemorrhagic shock rats

AIM: To investigate the effect of ischemic preconditioning (IPC) on vascular reactivity and calcium sensitivity during hemorrhagic shock. METHODS: Appropriate method of IPC was selected by observing the effect of different strategies of IPC on the survival time and the survival rate in hemorrhagic shock rats. The effect of IPC on the pressor effect of norepinephrine (NE, 3 μg/kg) and the contractile response of superior mesenteric artery (SMA) to NE and calcium in vivo and in vitro were observed. RESULTS: Among 3 strategies of IPC, 3 cycles of abdominal aorta occlusion for 1 min and loosing for 5 min increased the survival time and 24 h survival rate significantly, which was superior to the other two IPC methods. In vivo, IPC significantly increased the pressor response to NE and the contractile response of SMA to NE (P<0.01). In vitro, IPC significantly improved the reactivity of SMA to NE and Ca²⁺. The E_max values of SMA to NE and Ca²⁺ in IPC group were significantly higher than that in shock control group (P<0.01). CONCLUSION: Ischemic preconditioning reverses Shock-induced vascular hyporeactivity via improving calcium sensitivity of the vasculatures.     

Mechanism of mesenteric lymph duct ligation against the renal insufficiency in hemorrhagic shock rats

AIM: To observe the effect of mesenteric lymph duct ligation on free radical and inflammatory mediator in serious hemorrhagic shock rats at different periods, and explore the mechanism of intestinal lymphatic pathway on renal insufficiency. METHODS: 78 male Wistar rats were divided into the sham group, shock group, and ligation group. The model of serious hemorrhagic shock was established in shock group, ligation group, and mesenteric lymph was blocked by ligating mesenteric lymph duct in ligation group after resuscitating. All rats were executed and kidneys were taken out for making homogenate of 10 percent to determine levels of MDA, SOD, NO, NOS, tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and myeloperoxidase (MPO) at time points after shock 90 min, after transfusion and resuscitate 0 h, 1 h, 3 h, 6 h, 12 h and 24 h. The expression of inducible nitric oxide synthase (iNOS) mRNA in kindey was detected by RT-PCR. RESULTS: The contents of MDA, NO, NOS, TNF-α, IL-6, MPO and iNOS expressions in renal homogenate of shock group were increased after transfusion and resuscitation, and were higher at 6 h and 12 h, and was significantly higher than that in sham group. The acvitity of SOD was significantly lower than that in sham group (P<0.01, P<0.05). The contents of MDA, NO, NOS, TNF-α, IL-6, MPO and iNOS expression in renal homogenate of ligation group after transfusion and resuscitation 6 h, 12 h and 24 h were significantly lower than those in shock group at same points, and the SOD activity was higher (P<0.01, P<0.05). CONCLUSION: The results demonstrate that the ligation of mesenteric lymph duct can antagonise the development of renal failure in serious hemorrhagic shock rats, and its mechanism might relate to reduce the PMN sequestration, decrease the levels of TNF-α and IL-6, inhibit NO production and expression of iNOS mRNA, suppress the release of free radical and consumption of SOD.     

Effect of non-invasive ischemic preconditioning on nitric oxide/endothelin-1 imbalance and gas exchange impairment following limb ischemia reperfusion: a clinical study

AIM: To investigate the effects of non-invasive ischemic preconditioning on nitric oxide (NO)/endothelin-1 (ET-1) imbalance and gas exchange impairment following limb ischemia reperfusion in patients undergoing unilateral lower extremity surgery with tourniquet. METHODS: Twenty-seven patients aged 25-65 years, whose tourniquets duration varied from 1 h to 1.5 h and matched American Society of Anesthesiologists Physical Status Ⅰ-Ⅱ, were randomized into two groups: a control group (n=14) and a ischemic preconditioning group (IPC group, n=13) in which patients received three cycles of 5 min of ischemia/5 min of reperfusion before tourniquet inflation. Radial arterial blood gas, plasma malondialdehyde (MDA) and NO, serum ET-1 and interleukin-6 (IL-6) were measured just before tourniquet inflation(T0), 1 h after inflation(T1), and 0.5 h(T2), 2 h(T3), 6 h(T4), 24 h(T5) after tourniquet deflation. Meanwhile NO/ET-1 ratio, alveolar-arterial oxygen gradient (PA-aDO2) and intrapulmonary shunt (Qs/Qt) were calculated. RESULTS: In control group, arterial partial pressure of oxygen (PaO2) were decreased, while PA-aDO2 and Qs/Qt were increased significantly at T4 compared to the baselines at T0 (P<0.01). Plasma NO levels and NO/ET-1 ratios decreased gradually after tourniquets deflation and statistical significances were observed at T3 (P<0.01) with a valley at T4 (P<0.01) and recovered to baselines at T5. Serum ET-1, IL-6 and plasma MDA began to increase remarkably after T3 (P<0.05 or P<0.01), peaked at T4 and dropped slightly at T5. The changes above-mentioned could be well attenuated by the application of IPC (P<0.05 or P<0.01) except PaO2 (P>0.05). CONCLUSION: Clinical application of unilateral tourniquet within safe time limit (1.5 h) may lead to limb ischemia reperfusion and further pulmonary gas exchange impairment, which could be partially attenuated by the application of IPC via alleviating NO/ET-1 imbalance.