Objective To investigate the molecular mechanism of multiple cellular factors expressed shortly after ischemia reperfusion (IR) injury from the pathway of nuclear factor kappa B (NF κB). Methods The isolated heart models were established and sixty six rats were randomly divided into experimental group and control group. The deoxyribonucleic acid (DNA) binding activities of NF κB, the inhibitory kappa B (IκBα) levels in cytoplasm and tumor necrosis factor α (TNF α) messenger ribonucleic acid (mRNA) expressions were determined after 5, 15 min ischemia in experimental group, both after 0, 5, 15, 30 min ischemia and concomitantly 5, 15, 30, 45, 60 min reperfusion in control group. Results Augment of DNA binding activities of NF κB and reduction of IκBα in cytoplasm shortly after ischemia results were observed in control group. The level of IκBα was restored after reperfusion, the DNA binding activities of NF κB was further augmented. DNA binding activities of NF κB and TNF α mRNA expressions were lower in experimental group than those in control group. Conclusions NF κB in IR myocardium is activated by two different pathways: p65 p50 heterodimers and p50 p50 homodimers. In addition, the results suggest that early activation of NF κB induced by ischemia in the myocardium could be a signal mechanism for controlling and regulating immediate gene expressions during ischemia reperfusion.
Objective To investigate the effect of N-acetylcysteine (NAC) on the apoptosis during myocardial ischemia reperfusion injury in rats’ heart transplantation, and to explore the possible role of NAC in myocardial apoptosis. Methods Sixty healthy male Lewis rats (weighing, 200-220 g) were randomly divided into 3 groups, 20 rats each group (10 donors and 10 recipients). In control group, 1 mL normal saline was infused via inferior vena cava at 30 minutes before donor harvesting; in donor preconditioning group, NAC (300 mg/kg) was infused via inferior vena cava at 30 minutes before donor harvesting, but no treatment in recipients; and in recipient preconditioning group, NAC (300 mg/kg) was infused via inferior vena cava at 30 minutes before recipient transplantation, but no treatment in donors. Heart transplantation was established in each group. Blood was drawn at 6 and 24 hours after reperfusion for analysis of aspartate aminotransferase (AST), alanine aminotransferase (ALT), lactate dehydrogenase (LDH) as markers of graft injury; myocardial tissue was harvested to determine the superoxide dismutase (SOD) and lipid hydroperoxide (LPO) activity at 24 hours after reperfusion and to observe the histology and ultrastructural changes. Graft active Caspase-3 protein expression was measured by immunohistochemistry staining, and apoptosis index (AI) was calculated by TUNEL. Results The heart transplantation operation was successfully completed in all groups, and the rats survived to the end of the experiment. The serum levels of AST, ALT, and LDH in donor and recipient preconditioning groups were significantly lower than those in control group at 6 hours after reperfusion (P lt; 0.05); the levels of AST and ALT in donor preconditioning group and the levels of AST and LDH in recipient preconditioning group were significantly lower than those in control group at 24 hours (P lt; 0.05); and no significant difference was found between donor and recipient perconditioning groups (P gt; 0.05). The levels of AST, ALT, and LDH at 24 hours were significantly lower than those at 6 hours in each group (P lt; 0.05) except the level of ALT in recipient preconditioning group (P gt; 0.05). SOD activity and SOD/LPO in donor and recipient preconditioning groups were significantly higher than those in control group (P lt; 0.05), but no significant difference between donor and recipient preconditioning groups (P gt; 0.05); there was no significant difference in LPO activity among 3 groups (P gt; 0.05). Histological staining and transmission electron microscope showed that myocardial injury in recipient preconditioning group was obviously lighter than that in donor preconditioning group and control group. Active Caspase-3 in recipient pretreatment group was significantly higher than that in donor preconditioning group and control group (P lt; 0.05). AI of donor and recipient preconditioning groups was significantly lower than that of control group (P lt; 0.05), but no significant difference was found between donor and recipient preconditioning groups (P gt; 0.05). Conclusion NAC can relieve ischemia reperfusion injury in rats’ heart transplantation by improving myocardial SOD content, and reducing active Caspase-3 activity and AI, which has a protective effect on myocardial cell of donor heart.
OBJECTIVE: To observe the changes of heme oxygenase-1 (HO-1) expression in the skeletal muscle after ischemia-reperfusion of hind limb in rats. METHODS: A model of hind limb ischemia was made by clamping femoral artery with a microvascular clip. Soleus muscle was obtained from the animals received sham operation, 4 h ischemia without reperfusion and 2 h, 4 h, 8 h, 16 h, 24 h reperfusion after 4 h ischemia. Soleus histology and malondialdehyde (MDA) content were measured. The levels of HO-1 mRNA and protein were measured in different time by Northern blotting, Western blotting and immunohistochemistry technique. RESULTS: After ischemia-reperfusion of limb, HO-1 mRNA increased at the 2nd hour, reached a peak at the 8th hour, and returned toward baseline at the 24th hour. The change of protein level was essentially in agreement with that of mRNA. Immunohistochemical results showed that HO-1 expressed primarily in skeletal muscle cytoplasma. There were no positive signals of mRNA and protein in sham group and in ischemia group. After limb reperfusion, MDA contents in the soleus muscle increased significantly when compared with that in the sham group (P lt; 0.05). MDA content of the 8th after reperfusion decreased significantly when compared with that of the 4 h after reperfusion (P lt; 0.05). CONCLUSION: Ischemia-reperfusion can induce HO-1 expression in skeletal muscle in rats, which may provide protection for injured tissue.
Objective To summarize the function of Kupffer cell for the ischemia reperfusion injury after liver’s transplatation. Methods The literatures which about the function of Kupffer cell for the ischemia reperfusion injury after liver’s transplatation were reviewed. Results Kupffer cells are the resident macrophages of the liver, which can be activated to generate a range of inflammatory mediators, including cytokines, reactive oxygen intermediates, chemokines, and other factors to startup the ischemia reperfusion injury （IRI）, and to cause the liver graft dysfunction. On the other hand, Kupffer cells can protect the ischemia reperfusion injury by release NO and HO-1. The CO, which is the byproduct of heme degradation by the heme oxygenases （HO-1），has the same function for IRI. Conclusions The Kupffer cells have bidirectional function for the ischemia reperfusion injury of liver’s transpatation. Thus, how to decrease the harmful factors and up-regulate the beneficial substances by Kupffer cells will be the key points in preventing IRI after liver transplantation in future.
Objective To study the interaction and mechanism of prostaglandin I2 (PGI2) receptor/thromboxane A2 (TxA2) receptor (IP/TP) and cyclooxygenase-2 (COX-2) in ischemia reperfusion injury after liver transplantation of rat. Methods Rats were randomly divided into three groups: control group （n=16), orthotropic liver transplantation group （n=32) and nimesulide intervention group （n=32). The samples were obtained at 3 h, 6 h, 12 h and 24 h after operation. The expressions of COX-2, IP and TP mRNA were detected by RT-PCR. Immunohistochemistry was used to detect the localization and expression of COX-2. Hematoxylin Eosin staining was used to classify the injury extent of liver. Serum ALT and AST levels were detected to evaluate the changes of liver enzyme. Results COX-2 protein expression detected by immunohistochemistry in orthotropic liver transplantation group mainly distributed in the district of liver sinusoidal endothelial cells, liver cells and macrophage cells, which was significantly higher than control group and nimesulide intervention group. Expressions of IP mRNA, TP mRNA and COX-2 mRNA in the orthotropic liver transplantation group were significantly increased than those in control group (P＜0.05), and the ratio of IP/TP increased (P＜0.05). Expressions of IP mRNA and TP mRNA in nimesulide intervention group were significantly lower than that in the orthotropic liver transplantation group at 6 h and 12 h after operation (P＜0.05), and the ratio of IP/TP decreased at 3 h, 6 h and 24 h after operation (P＜0.05). The expression of COX-2 mRNA in nimesulide intervention group was significantly lower than that in the orthotropic liver transplantation group at 6 h, 12 h and 24 h after operation. In orthotropic liver transplantation group liver injury was obvious by HE staining, and more severve than that in nimesulide intervention group. Serum AST (each time) and ALT (3 h, 6 h and 12 h) levels in the orthotropic liver transplantation group were significantly higher than that in control group and nimesulide intervention group (P＜0.05) and peaked at 6 h after operation. Conclusion The balance of IP/TP takes part in and plays an important role in the ischemia reperfusion injury of liver transplantation. Changing imbalance of IP/TP may reduce liver transplantation ischemia reperfusion injury by inhibiting COX-2 expression.
ObjectiveTo explore the effect of hypertonic saline (HTS) pretreatment on levels of nitric oxide (NO) and endothelin-1(ET-1) and their correlation in hepatic ischemia reperfusion (HIR) injury in rats. MethodsThe HIR injury models were made by using Pringle, s maneuver in 45 healthy adult male Sprague-Dawley rats, which were randomly divided into three groups (n=15):sham operation (SO) group, HIR group, and HTS group. The animals were killed at 1, 6, and 24 h after reperfusion. The levels of serum NO and ET-1 were measured respectively, the correlation between NO level and ET-1 level at 6 h after reperfusion was analyzed. ResultsAt the time points of 1 h, 6 h, and 24 h after reperfusion, the serum NO levels in the HTS group and HIR group were all significantly lower than those in the SO group (P < 0.01), but the serum ET-1 levels were all significantly higher than those in the so group (P < 0.01). The serum NO levels at the time points of 1 h, 6 h, and 24 h in the HTS group were significantly higher than those at the same time in the HIR group (P < 0.01), but the serum ET-1 levels in the HTS group were significantly lower than those in the HIR group (P < 0.01). At all the time points, every detected goal had more marked level at the time point of 6 h after reperfusion. The NO level was negatively correlated with the ET-1 level. ConclusionsHTS could change levels of serum NO and ET-1 after HIR injury, and which has a negative correlation. Its mechanism might probably stimulate serum NO level and reduce the ET-1 level through some way so as to enable both dynamic balance to the benign development direction and achieve a protective effect.
ObjectiveTo investigate the effect of power-assisted intravascular shunt in replantation of amputated limbs of rabbits. MethodsEighty rabbits weighing 1.8-2.5 kg (male or female) were selected to establ ish the model of circular amputation at the hind groin, only femoral arteries and veins were completely preserved. After the femoral artery was clamped in 60 rabbits, the rabbits underwent power-assisted intravascular shunt with high-flow rate (group A, n=20), powerassisted intravascular shunt with low-flow rate (group B, n=20), and no power-assisted intravascular shunt (group C, n=20) to reconstruct blood supply; the femoral artery was not clamped in another 20 rabbits of sham group (group D). Before and after intravascular shunt (1, 3, 6, and 12 hours), the malondialdehyde (MDA), lactate dehydrogenase (LDH), and creatine kinase (CK) of the serum were determined. The myeloperoxidase (MPO), MDA, and wet to dry weight ratio (W/D ratio) of the gastrocnemius muscle were measured, and the thrombogenesis and survival rate of limb were observed. ResultsBefore intravascular shunt, MDA, LDH, and CK of the serum and MPO, MDA, and W/D ratio of the muscle showed no significant difference among 4 groups (P>0.05). At each time point after intravascular shunt, no significant difference was found in all indexes between groups A and D (P>0.05); the indexes of groups B and C were significantly higher than those of groups A and D (P<0.05); the values were the highest in group C (P<0.05), and reached the peak at 12 hours. All limbs of group A survived with low thrombosis rate, and less limbs could survive with high thrombosis rate in group C. ConclusionThe power-assisted intravascular shunt with high-flow rate can effective ensure the blood supply of the amputated limbs of rabbits with lower limb injury and higher survival rate of amputated limbs after replantation.