Role of IRAK-4 activity in inhibitory effects of lipopolysaccharide pretreatment on hepatic ischemia/reperfusion injury

AIM:To observe the changes of interleukin-1 receptor associated kinase-4 (IRAK-4) in ischemia/reperfusion (I/R) liver pretreated with lipopolysaccharide (LPS) and to explore the protective mechanisms of LPS pretreatment against hepatic I/R injury. METHODS:Male Sprague-Dawley rats, weighing 240-280 g, were divided into three groups:control, ischemia/reperfusion group (I/R group) and LPS-pretreated group (LPS group). On the first day, LPS group received 0.1 mg/kg LPS via the tail vein, followed by 0.5 mg/kg on the 2nd, 3rd, 4th and 5th day. I/R group received the equivalent volumes (0.5 mL) of sterile PBS. Experiments of I/R injury was induced by temporary ischemia of the left lateral liver lobe for 90 min followed by 3 h reperfusion on 2 days after the last LPS treatment. At 0 min, 60 min and 180 min after reperfusion, the expression of IRAK-4 gene and protein level were determined by RT-PCR and Western blotting. The activity of NF-κB and the serum TNF-α level were also detected by ELISA. RESULTS:Although the level of IRAK-4 gene and protein were higher in the LPS group than that in I/R group and control group (P<0.01), no difference of the activities of NF-κB and the TNF-α level was observed between the LPS group and I/R group (P>0.05) at 0 min after reperfusion. However, all those indexes were evidently lower in the LPS group than those in I/R group (P<0.01) at 60 min and 180 min after reperfusion. CONCLUSION:This data suggests that the protective effects induced by LPS pretreatment against hepatic I/R injury may be via down-regulation of IRAK-4 expression.     

Establishment and evaluation of rat heart ischemia-reperfusion injury model in vivo

AIM: To establish and evaluate a rat model of heart ischemia-reperfusion injury in vivo. METHODS: Seventy-two male Sprague-Dawley rats weighing(250±50)g were randomly divided into sham operation group(sham), ischemia-reperfusion group(I/R) and normal group. The animals were anesthetized and heparinized. Myocardial ischemia-reperfusion was induced by ligating the left anterior descending coronary artery with "U-shape tube" for 35 min followed by 120 min or 240 min reperfusion in vivo. The heart infarct size was measured by triphenyltetrazolium chloride(TTC) staining. The myocardial cell apoptotic index was determined by the method of terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling(TUNEL). Immunohistochemical method was used to detect the expression of Bcl-2 and Bax in rat ischemia myocardium. The blood level of MB isoenzyme of creatine kinase(CK-MB),cardiac troponin I(cTnI),nitric oxide(NO),malondialdehyde(MDA), total superoxide dismutase(T-SOD)and glutathione peroxidase(GSH-Px) were detected after reperfusion for 2 h and 4 h. RESULTS: Compared with normal group and sham group, there were obvious changes of ST-T segment and Q wave in the electrocardiogram of I/R group. The blood level of CK-MB, cTnI, NO, MDA and GSH-Px in I/R group increased(P<0.05,P<0.01) after reperfusion for 2 h and 4 h, and the blood level of T-SOD in I/R group after reperfusion for 2 h and 4 h also increased(P<0.05). The heart infarct size in I/R group was the largest as compared to other groups. Among these groups, the apoptotic index of I/R group was the highest and the Bcl-2/Bax ratio in I/R group decreased(P<0.01).CONCLUSION: The rat model of heart ischemia-reperfusion injury in vivo can be successfully established with the "U-shape tube". There are obviously changes of heart infarct size, blood level of CK-MB, cTnI, NO, MDA, T-SOD and GSH-Px, myocardial apoptotic index and Bcl-2/Bax ratio between I/R rats and control animals.     

Pretreatment of hypertonic saline attenuates the hepatic ischemia reperfusion injury induced by neutrophils

AIM: To explore the effect of the pretreatment of hypertonic saline (HTS) in hepatic ischemia reperfusion (I/R) injury.METHODS: The rats were divided into sham group (sham group), ischemia reperfusion group (IR group) and pretreatment of hypertonic saline group (HTS group). Partial hepatic ischemia reperfusion model was used. The rats were sacrificed at the time of 1 h, 3 h, 6 h, 12 h and 24 h after reperfusion in each group, respectively. Blood samples were obtained to examine ALT. The expression of the CD11b/CD18 (Mac-1) on the neutrophils was analyzed by flow cytometry. RT-PCR and Western blotting were used to examine the expression of intercellular adhesion molecule-1 (ICAM-1) in livers and chromatometry was performed to detect the activity of myeloperoxidase (MPO) in livers. The morphology of hepatocytes and the structure of sinusoid were observed by histological examinations. RESULTS: ① HTS pretreatment decreased the level of ALT at the time points of 3 h, 6 h and 12 h after reperfusion (P<0.05). ② Mac-1 expression in HTS group was lower at 6 h and 12 h after reperfusion compared with IR group (P<0.05). ③ MPO activity in HTS group was lower at 6 h, 12 h and 24 h compared with IR group (P<0.05). ④ RT-PCR and Western blotting analysis indicated that the pretreatment of HTS inhibited the expression of ICAM-1 in livers after reperfusion. ⑤ Moderate hepatocyte swelling and few neutrophil infiltration were observed in HTS group.CONCLUSION: Pretreatment with HTS has the effect on hepatic ischemia reperfusion injury by inhibiting the expression of Mac-1 on circulating neutrophils and the expression of ICAM-1 in the liver.     

Effects of ethane 1,2-dimethanesulfonate preconditioning on renal ischemia/reperfusion injury in male rats

AIM: To investigate the effects of ethane 1,2-dimethanesulfonate (EDS) preconditioning on renal ischemia/reperfusion (I/R) injury in male Sprague-Dawley (SD) rats. METHODS: Male SD rats (n=48) were randomly assigned to 6 groups: blank, sham, I/R, EDS+I/R, EDS+testosterone (TST)+I/R, and castration (Cast)+I/R. The renal pedicles were bilaterally occluded with a microvascular clamp for 45 min to establish renal I/R-induced injury model. Bilateral orchiectomy was conducted 2 weeks before surgery. EDS (75 mg/kg) was intraperitoneally injected 5 d before operation. Blood samples were collected 24 h after reperfusion from the vena orbitalis posterior plexus. Luteinizing hormone (LH), TST, serum creatinine (SCr), blood urea nitrogen (BUN), and kidney injury molecule-1 (KIM-1) were detected. The renal tissues were harvested to measure the level of TNF-α and the expression of Fas mRNA and caspase-3 protein. RESULTS: Serum TST levels in EDS+I/R group and Cast+I/R group were below the minimum detectable threshold. Compared with other groups, the rats in EDS+I/R group and Cast+I/R group had higher levels of SCr, BUN and KIM-1 (P<0.05). SCr and BUN levels showed no significant difference between EDS+I/R group and Cast+I/R group (P>0.05), but KIM-1 level in EDS+I/R group was lower than that in Cast+I/R group (P<0.05). After reperfusion for 24 h, the levels of TST and LH in EDS+I/R group, Cast+I/R group and EDS+TST+I/R group were lower than those 1 h before operation (P<0.05). Compared with Cast+I/R and I/R group, the TNF-α level and expression of Fas mRNA and caspase-3 protein were significantly decreased in EDS+I/R group (P<0.05). CONCLUSION: EDS preconditioning substantially reduces the serum TST level, thus attenuating I/R-induced acute renal injury. TNF-α-induced Fas/FasL pathway may be involved in this process.     

Significance and changes of serum interleukin-8 and 10 in human myocardium ischemic preconditioning

AIM: To explore the mechanism of myocardium protection after ischemia/reperfusion (I/R) injury by preconditioning with ischemia in human. METHODS: Thirty-six patients underwent valve replacement were divided into ischemic preconditioning group (IP group, 20 cases) and non-ischemic preconditioning group (control group, 16 cases) according to whether they were given single cycle reperfusion before cardioplegia or not. Serum levels of interleukin-8 and 10 were measured with ELISA. Expressions of myocardial Bcl-2 and caspase-3 were analyzed. RESULTS: The inflammatory factors IL-8 and IL-10 increased to the highest level in serum at 6 h after declamping and recovered to normal level on 5 d after declamping. On 6 h, 1 d and 2 d after declamping, serum level of IL-8 was significantly lower in IP group than that in control group (P<0.05), but serum level of IL-10 was higher in IP group (P<0.05). Expression of myocardial Bcl-2 and caspase-3 increased in both groups after reperfusion, and Bcl-2 was lower in the control group than that in IP group while the level of caspase-3 was higher (P<0.05). Expression of myocardial Bcl-2 had positive correlation with IL-10 and negative correlation with IL-8. CONCLUSION: Ischemic preconditioning has the effect of protection of human myocardial cells after ischemia/reperfusion injury through decreasing systemic inflammatory response following ischemia reperfusion injury.     

Effects of electrical stimulation of vagus nerve on gut injury following intestinal ischemia-reperfusion in rats

AIM:To investigate the effects of electrical stimulation of vagus nerve on gut injury following intestinal ischemia-reperfusion in rats. METHODS:30 adult male Wistar rats subjected to bilateral cervical vagotomy were randomly divided into three groups (n=10 per group):(1) Intestinal ischemia-reperfusion group (group I/R):laparotomy and I/R induced by clamping arteria mesenterica superior for 1 h followed by reperfusion for 2 h. (2) Vagus nerve stimulation group (group VNS):laparotomy, I/R and electric stimulation with pulse train of constant amplitude 5V, pulse width 2 ms and frequency 1 Hz at the left caudal vagus ends for 20 minutes before and after occlusion. (3) Sham control group (group SC):sham operation and sham stimulation. Carotid artery was cannulated for mean arterial pressure (MAP) monitoring. A strip of small intestine was taken from distal end of ileum for light microscopic (LM) and transient electron microscopic (TEM) examination at the time of 2 h after reperfusion. Improved Chiu’s scale was used to quantitatively assay the damage degree. The levels of malondialdehyde (MDA) and TNF-α in plasma were detected. RESULTS:MAP in every group kept steady during ischemia, but decreased gradually with the prolongation in the time of reperfusion. MAP decreased more dramaticly in group I/R than that in group VNS (P<0.05). Histological changes by LM and TEM were more significant in group I/R than those in group VNS and SC. The improved Chiu’s scale in group VNS was obviously slighter than that in group I/R (P<0.05). In group I/R, the levels of MDA and TNF-α in plasma were significantly increased compared with group VNS and SC (P<0.01, P<0.05). MDA level in group VNS was significantly higher than that in group SC, but there was no significant difference in TNF-α between the two groups. CONCLUSION:Electrical stimulation of vagus nerves can lessen pathological changes of intestinal mucosa induced by I/R and improve BP during reperfusion, which may be related to reducing lipid peroxidation, and down-regulating TNF-α synthesis.     

Influence of high-mobility group box 1 on proliferation of neural stem cell in pre-infarction cortex of focal cerebral ischemia/reperfusion model rats

AIM：To investigate the influence of high-mobility group box 1 (HMGB1) on the proliferation of neural stem cells in peri-infarction cortex of focal cerebral ischemia/reperfusion model rats. METHODS：Male SD rats (n=48) were randomly divided into sham group, ischemia/reperfusion (I/R) group, RNA interference group and negative interference group. The rat middle cerebral artery was blocked to establish focal cerebral I/R model (ischemia for 1 h and reperfusion for 7 d). Lentivirus vector of HMGB1 shRNA was used to suppress the HMGB1 protein expression in the rat brain. The effect of RNA interference was evaluated by the methods of double-immunofluorescence labeling of HMGB1/GFAP and Western blotting. The proliferation of neural stem cells in the peri-infarction cortex was assessed by double labeling of BrdU/nestin. RESULTS：The protein expression of HMGB1 in I/R group was much higher than those in sham group (P<0.05). RNA interference effectively inhibited the HMGB1 expression (P<0.05). Double labeled BrdU/nestin positive cells in I/R group were more than that in sham group (P<0.05). The double labeled BrdU/nestin positive cells were significantly decreased in RNA interference group (P<0.05). CONCLUSION：Focal cerebral ischemia/reperfusion injury promotes the proliferation of neural stem cells in peri-infarction cortex by increasing HMGB1 protein level.     

Inhibitory effect of ginkgo-dipyridamole injection on ischemia/reperfusion injury in rat hearts in vitro

AIM：To investigate the effect of ginkgo-dipyridamole injection (GD) on ischemia/reperfusion (I/R) injury in rat hearts in vitro and its possible mechanism. METHODS：Forty male Sprague-Dawley rats were randomly divided into 5 groups (n=8): normal control (NC) group, I/R group, ischemic preconditioning (IPC)+I/R group, GD+I/R group and GD+LaCl3+I/R group. Cardiac function indexes, including heart rate (HR), left ventricular systolic pressure (LVSP) and the maximal rise/fall rate of left ventricular pressure (±dp/dtmax), were detected at 5 time points, including stabilizing point, 30 min after ischemia, and 5, 30 and 60 min after reperfusion. The activity of lactate dehydrogenase (LDH) and creatine kinase (CK) in coronary effluent at the five time points was assayed. The concentration of Ca2+ and the content of α-ketoglutarate dehydrogenase (α-OGDH) in myocardial mitochondria were determined at the end of the whole experiment. RESULTS：Compared with I/R group, the cardiac function indexes in IPC+I/R and GD+I/R groups were improved at the reperfusion period (P<0.05), the activity of LDH and CK in coronary effluent and the concentration of Ca2+ in mitochondria were significant reduced (P<0.01), and the content of α-OGDH was increased (P<0.05). However, the protective effect of GD was inhibited by LaCl3 (P<0.05). CONCLUSION：GD protects rat hearts against I/R injury by inhibiting calcium overload and improving mitochondrial enzyme activity to stabilize mitochondrial energy metabolism.     

Effects of ischemic preconditioning on hepatocyte apoptosis induced by hepatic ischemia-reperfusion in cirrhotic rats

AIM:To investigate the protective effect of ischemic preconditioning (IPC) on hepatic ischemia-reperfusion(I/R) injury in cirrhotic rats and its possible mechanism. METHODS:Hepatic I/R was induced by Pringle maneuver. The cirrhotic rats were randomized into three groups: Group A: before 30 min of ischemia, a short period of 5 min ischemia and 5 min reperfusion were given; Group B: before 30 min of ischemia, a short period of 10 min ischemia and 10 min of reperfusion were given; Group C: 30 min ischemia only. The serum alanine transferase (ALT), hepatic Fas-mRNA, caspase-3 activity and hepatocyte apoptosis were analyzed. RESULTS:The 7-day survival rate in the group A and B were 100%, respectively. However, it was only 62.5% in the group C. After 6 h of reperfusion, the ALT levels in both group A and B were significantly lower than that of in group C, P＜0.01. The ALT level of group A was also lower than that of group B, P＜0.01. The hepatic Fas-mRNA expression, caspase-3 activity and apoptotic hepatocyte in group A were significantly lower than those of in group C, P＜0.01. CONCLUSIONS:IPC has significant protective effect against hepatic I/R injury. An IPC with 5 min of ischemia and 5 min of reperfusion has the maximal protective effect. The protective mechanism of IPC against hepatic I/R injury is via down-regulation of Fas-mRNA expression, inhibiting caspase-3 activity and subsequently inhibiting hepatocyte apoptosis.     

Changes of gut mucosa antioxidant system and liver,renal function in rats after intestinal ischemia-reperfusion injury

AIM: To investigate the changes of the gut mucosa antioxidant system and liver, renal functions during rat intestinal ischemia-reperfusion injury. METHODS: 30 male Wistar rats underwent 45 min of intestinal ischemia by clamping the superior mesenteric artery followed by reperfusion. The levels of malondialdehyde (MDA), glutathione (GSH), the activities of antioxidant enzymes in the gut mucosa including catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), glutathione S- transferase (GST) activity and serum ALT, AST, BUN, Cr were assayed at 2, 4, 8, 12 and 24 h after reperfusion. RESULTS: The levels of MDA and GSH in the gut mucosa increased and decreased significantly at 2 h of reperfusion, respectively (P<0.05). MDA was still lower than sham at 24 h of reperfusion (P<0.05), while GSH decreased to 40% of sham at 4 h of reperfusion (P<0.01) but returned to the level of control at 12 h. The activities of CAT, SOD and GSH-Px did not show significant changes in rat after intestinal ischemia reperfusion. GST decreased 39% at 2 h of reperfusion compared with the sham group and decreased to 56% of sham at 4 h (P<0.05), but returned to the level of control at 12 h after reperfusion. Serum ALT, AST, BUN and Cr increased significantly at 2 h of reperfusion (P<0.05) and increased 208%, 100%, 103%, 41% compared with control at 4 h of reperfusion (P<0.01). However, at 24 h of reperfusion, they returned to normal. CONCLUSION: Intestinal ischemia/reperfusion diminishes GSH level and GST activity, increases MDA level and causes liver and renal reversible damages.     

Effects of 11, 12-EET and ischemic preconditioning on phosphorylated ERK and p38 MAPK during ischemia and reperfusion in rat myocardium

AIM: In order to study the relationship between the ERK and p38 MAPK activation and the protection of 11, 12-epoxyeicosatrienoic acid (11, 12-EET) and ischemia preconditioning (IP), the effects of 11, 12-EET and ischemic preconditioning on phosphorylated ERK and p38 MAPK during ischemia and reperfusion in rat myocardium were examined. METHODS: The rat heart was subjected to ischemia for 5 min by ligating the left anterior descending coronary artery followed by reperfusion for 5 min (two times) to undergo ischemia preconditioning. The rats were divided into 5 groups: (1) control; (2) sham group; (3) ischemia/reperfusion (I/R) group, in which the rat heart suffered from 60 min ischemia followed by 30 min reperfusion; (4) IP plus I/R group; (5) EET plus I/R group, in which 6.28×10^-8 mol/L 11, 12-EET was injected intravenously 20 min before I/R. The heart function was examined, and phosphorylated ERK and p38 MAPK were detected by Western blot. RESULTS: At 30 min reperfusion, +dp/dt_max, -dp/dt_max and LVDP decreased significantly in I/R group compared with sham group, IP plus I/R group and EET plus I/R group; Phosphorylated ERK1/2 level was higher in I/R group than sham group, but was lower in I/R group than IP plus I/R group and EET plus I/R group; Phosphorylated p38 MAPK level was lower in control, sham, IP plus I/R and EET plus I/R group than I/R group. CONCLUSION: 11,12-EET protects rat heart against ischemia/reperfusion injury, the mechanism may be related to activation of ERK1/2 and inhibition of p38 MAPK.     

Effects of glutamine pretreatment on intestinal ischemia-reperfusion injury in rats

AIM：To determine the effects of glutamine (Gln) pretreatment on intestinal ischemia-reperfusion (I/R) injury in the rats. METHODS：Thirty male Wistar rats were randomly divided into 3 groups (n=10): sham group, I/R group and Gln pretreatment group. The rats in Gln pretreatment group were pretreated with 1 g·kg-1·d-1 Gln by orogastric route for 7 d, the rats in the other 2 groups were pretreated with normal saline. Intestinal I/R was induced by 30-min occlusion of the superior mesenteric artery followed by 24 h of reperfusion. After the operation, the plasma endotoxin, serum D-lactic acid, superoxide dismutase (SOD) and malondialdehyde (MDA) levels were measured. The intestinal mucosal injury was observed with HE staining and evaluated using Chius scoring. RESULTS：Serum D-lactic acid, endotoxin level, MDA level and Chiu's score in I/R group were significantly higher than those in sham group and Gln group (all P<0.05). Serum SOD activity was significantly lower than that in sham group and Gln group (P<0.05). CONCLUSION：Glutamine has a protective effect on the intestines during ischemia-reperfusion injury. The mechanism may be related to oxidative stress response.     

Ischemic postconditioning protects skeletal muscle from ischemia/reperfusion injury through improving microcirculation in rat hind limb

AIM: To investigate the effect of ischemic postconditioning (I-postC) on ischemia/reperfusion (I/R) injury of skeletal muscle and its microvasculatory mechanisms in rat hind limb. METHODS: Healthy male Wistar rats were randomly divided into three groups as follows (n=16): I/R group, I-postC group and ischemic preconditioning (IPC) group. I/R of right hind limb was induced by clamping right femoral artery. The effects of I-postC on oxygen saturation of hemoglobin (StO2), blood flow, microcirculation, hemodynamics, Wet/Dry ratio (W/D) and ultramicrostructure of skeletal muscle were detected on 12 h or 24 h after reperfusion. RESULTS: (1) Compared with I/R group, 12 h after reperfusion, W/D from I-postC group decreased 6.7% (P<0.05) and myofibrillae lysis of skeletal muscle and vacuolation of mitochondria attenuated than that in I/R group. 24 h after reperfusion,-dp/dtmax in I-postC group increased 10.46% (P<0.05). (2) It was also found that 24 h after reperfusion, blood velocity in venule increased 11.3% in I-postC group compared with that in I/R group (P<0.05), and internal diameter of arteriole and venule increased 39.8% and 24.1% compared with that in I/R group, respectively (P<0.01). No significant difference between I-postC and IPC groups (P>0.05) was found. CONCLUSION: I-postC protects the skeletal muscle against reperfusion injury by improving microcirculation.     

Effect of ischemic post-conditioning on Egr-1 and IL-1β expression in ischemia-reperfusion injured lung in rats

AIM: To investigate the protective effects of ischemic post-conditioning on the expression of early growth response factor 1 (Egr-1) and interleukin-1β(IL-1β) in ischemia-reperfusion injured lung in rats. METHODS: The model of lung ischemia-reperfusion injury was established in 24 rats and the rats were randomly allocated to 3 different groups (n=8 in each group): (1) sham group: only sham operation (thoracotomy) and no ischemia for 3 h; (2)ischemia-reperfusion group (I/R group): interruption of pulmonary perfusion and ventilation for 1 h followed by reperfusion for 2 h; (3) ischemic post-conditioning group (IPostC group): ischemic post-conditioning (5 min of reperfusion and 5 min of ischemia for 3 times) between the end of ischemia and the beginning of the reperfusion followed by reperfusion for 1.5 h. The lung tissues (prepared to small pieces of about 20 mg) were collected and homogenized at the end of the experiment. The concentration of myeloperoxidase (MPO) in the homogenate was determined. The wet to dry weight ratio (W/D) of the lung tissues was also measured at the end of reperfusion. The pathological changes of the lung tissues were observed under light microscope after reperfusion. The mRNA expression of Egr-1 and IL-1β in the lung tissues was detected by RT-PCR. RESULTS: Compared with sham group, the mRNA expression of Egr-1 and IL-1β, the levels of MPO and W/D were significantly increased in I/R group (P<0.05). The inflammatory responses of the lungs in I/R group were significantly severer than those in sham group. Compared with I/R group, the mRNA expression of Egr-1 and IL-1β, the levels of MPO and W/D in IPostC group were significantly decreased (P<0.05). The inflammatory responses of the lungs in IPostC group were also significantly attenuated. CONCLUSION: Ischemic post-conditioning significantly reduces ischemic reperfusion injury of the lung by inhibiting the expression of Egr-1 and IL-1β.     

Correlation of peripheral leukocyte apoptosis insufficiency and intestinal injury following mesenteric ischemia/reperfusion in rats

AIM: To investigate the role of peripheral blood leukocytes and polymorphonuclear neutrophils (PMNs) on intestinal injury following mesenteric ischemia/reperfusion (IR) in rats. METHODS: Twenty adult, male Sprague-Dawley rats, weighing 200-230 g, were randomly divided into two groups. The control group (CON) consisting of 10 rats was subjected to laparotomy and separation of superior mesenteric artery (SMA) only. The ischemia/reperfusion (IR) group consisting of 10 rats, was subjected to laparotomy, followed by occlusion of the superior mesenteric artery (SMA) by an atraumatic microvascular clamp for 30 min. At the end of ischemic period in IR, the microvascular clamp was removed and the intestinal segment was reperfused for 60 min. The pathological changes of the ileal mucosal tissue were evaluated. The apoptosis of intestinal mucosal epithelial cells was examined by terminal deoxylnucleotidy-l transferase mediated-dUTP nick end-labeling (TUNEL). The enzymatic activity of casapse-3 in mucosal cells was determined using a colorimetric assay. The percentages of apoptotic peripheral blood leukocytes and PMNs were measured by flow cytometry using Annexin-V/PI double staining assay. The numbers of peripheral blood leukocytes in each animal was measured at baseline, 30 min of ischemia, and 30 min and 60 min of reperfusion. RESULTS: (1) Compared to CON group animals, the most severe mucosal injury was observed in IR group under optical microscope. (2) The number of apoptotic mucosal epithelia cells and enzymatic activity of caspase-3 were significantly higher in IR than those in CON group (P<0.05). (3) The percentages of apoptotic peripheral blood leukocytes and PMNs were significantly lower in IR group than those in CON group (P<0.05). The number of peripheral blood leukocytes in IR group was increased obviously after SMA closed for 30 min (P<0.05), and was higher following reperfusion (P<0.05), and significantly higher than that in CON group (P<0.05). (4) A negative correlation between the enzymatic activity of caspase-3 in the ileal mucosal tissue and the percentage of apoptotic peripheral blood leukocytes (r=-0.764, P<0.05), and of PMNs (r=-0.845, P<0.05) was found. A negative correlation between the apoptosis index of mucosal epithelial cells and the apoptotic percentage of PMNs (r=-0.638, P<0.05) was also observed. CONCLUSION: Apoptosis insufficiency and higher numbers of peripheral blood leukocytes are correlated with the mucosal cells injury following mesenteric ischemia/reperfusion (IR) in rats.     

Effects of astragaloside IV on autophagy in rats with cerebral ischemia/reperfusion injury

AIM: To investigate the effects of astragaloside IV (AS-IV) on autophagy in rats with cerebral ischemia/reperfusion (I/R) injury. METHODS: The focal cerebral ischemia/reperfusion of rat left middle cerebral artery occlusion (MCAO) was induced by suture method. Male SD rats (n=70) were randomly divided into sham operation group, I/R group, solvent control group, AS-IV group, AS-IV+autophagy inhibitor (3-methyladenine, 3-MA) group, 3-MA group and autophagy activator (rapamycin, Rapa) group. Except for sham operation group, the rats in other groups were subjected to ischemia for 2 h and reperfusion for 24 h. The rats with successful modeling were selected according to Zea Longa scoring criteria. The volume of cerebral infarction was measured by TTC staining. The morphological changes of nerve cells in the rats were observed with Nissl staining. The phenomenon of autophagy was observed under transmission electron microscope. The protein expression of beclin-1 and LC3-Ⅱ was determined by Western blot. RESULTS: No neurological deficit in sham operation group was observed, and the cerebral infarction was not found. Compared with sham operation group, obvious cerebral infarction was observed, the Nissl bodies were small in size and number and stained light, typical autophagosomes were observed, and the protein expression of beclin-1 and LC3-Ⅱ was increased in I/R group (P<0.05). Compared with I/R group, the volume of cerebral infarction was decreased obviously, neurological deficit restored significantly, and the number of autophagosomes and the protein expression of beclin-1 and LC3-Ⅱ were increased in AS-IV group and Rapa group (P<0.05). However, no significant difference between solvent control group and I/R group was observed (P>0.05). Compared with AS-IV group, the neurological deficit was serious, the volume of cerebral infarction and the number of autophagosomes were increased, while the expression of beclin-1 and LC3-Ⅱ was decreased in AS-IV+3-MA group and 3-MA group (P<0.05). CONCLUSION: Astragaloside IV may play an important role in atte-nuating cerebral ischemia/reperfusion injury by activating autophagy.     

Effect of pressure phase plane derived τ and K on evaluation of left ventricular diastolic function in isolated rat heart during ischemia/reperfusion

AIM: To analyze and compare the changes of pressure phase plane (PPP) derived τ and K on isolated rat heart during ischemia/reperfusion, and to explore the value of PPP derived τ and K for evaluation of left ventricular diastolic dysfunction. METHODS: LVEDP, -(dp/dt)_max, τ and K were measured and calculated during ischemia/reperfusion in Sprague-Dawley rat hearts. Meanwhile, the level of lactate dehydrogenase (LDH) in the coronary effluent was measured, and the ultrastructure changes in myocardium were observed under electron microscope. RESULTS: Compared with control group, τ increased and K reduced significantly in each ischemic group in a time dependent manner (P<0.05). With prolonged ischemia, τ was even higher and K was even lower (P<0.05). Compared with control group, except ischemia 15 min, LDH in other groups increased significantly at 10 min and 20 min after reperfusion (P<0.05). Compared with ischemia 30 min, LDH of ischemia 45 min and ischemia 60 min were even higher at 10 min and 20 min after reperfusion (P<0.05). With prolonged ischemia, the abnormal changes of the myocardial ultrastructure were observed. CONCLUSION: PPP derived τ and K may be promising indexes for quantitative assessment of left ventricular diastolic function on isolated rat heart during ischemia/reperfusion, and indication of the severity of ischemia/reperfusion injury.     

Effect of mild hypothermia on energy metabolism and hydroxyl radical production after cerebral ischemia/reperfusion in gerbils

AIM: To study the effect of mild hypothermia on energy metabolism and hydroxyl radical production as well as delayed neuronal death (DND) in hippocampus during cerebral ischemia/reperfusion in gerbils.METHODS: Forebrain ischemia was induced by occluding bilateral common carotid arteries with aneurysm clamps for 10 min in gerbils. The DND in hippocampal CA1 sector was assessed by histological examination, and hydroxyl radical, ATP (adenosine triphosphate), ADP (adenosine diphosphate),AMP (adenosine monophosphate) levels were determined by high performance liquid chromatography with electrochemical or ultraviolet detection. RESULTS: The number of survival neuronal in hippocampal CA1 sector in mild hypothermia + I/R group was more than that in I/R group after ischemia/reperfusion 96 h. The content of 2,3-DHBA (2,3- dihydroxybenzoic acid) in hippocampus in I/R group was much higher than those in sham operation and mild hypothermia + I/R group after reperfusion 6 h (P＜0.01), but there were no significant differences in 2,3-DHBA outputs among 3 groups 48 h and 96 h after reperfusion. There were no obvious differences in ATP, ADP, AMP level in hippocampus among 3 groups 6 h after reperfusion. However, the content of ATP,ADP,AMP in mild hypothermia + I/R group was much higher than those in I/R group 48 and 96 h after reperfusion (P＜0.01).CONCLUSION: Mild hypothermia can reduced DND by improving the cerebral energy metabolism during forebrain ischemia/reperfusion in gerbils.