Objective To investigate the protective mechanism of ischemic preconditioning on ischemic/reperfusion injury after pancreas transplantation in rats.Methods The model of diabetic SD rat was established. Twenty-four diabetic SD rats were randomly assigned to ischemic/reperfusion group (I/R group, n=6) and ischemic preconditioning group (IPC group, n=18). The rats in group IPC were averagely assigned to 3 sub-groups: group IPC_ 1 (5 min ischemic and 5 min reperfusion), IPC_ 2 (5 min ischemic and 5 min reperfusion twice) and IPC_ 3 (5 min ischemic and 5 min reperfusion thrice). Six normal SD rats whose abdomen was opened only served as control group, and they did not receive pancreas transplantation. I/R group and IPC group received pancreas transplantation. The superoxide dismutase (SOD) and myeloperoxidase (MPO) activity of grafts were monitored 2 h after reperfusion, the apoptotic cells in grafts were observed by TUNEL method, and the expression of Bcl-2 and Bax gene of the grafts was detected by Western blot.Results As compared with I/R group, the SIOD activity and the expression of Bcl-2 gene of grafts were significantly increased, while MPO activity, apoptotic index and the expression of the Bax gene in the grafts were markedly reduced in IPC group (P

BACKGROUND:Ischemia/reperfusion (IR)injury during the pancreas transplantation can cause numerous postoperative complications, among which,secondary pancreatitis can cause small intestinal mucosal injury and result in severe Consepuence.OBJECTIVE:To observe the protective effect of ischemic preconditioning (IPC) on small intestinal mucosal barrier after pancreas transplantation in rats.DESIGN:Randomized controlled animal trial.SETTING:Department of Gastrointestinal Surgery,Xijing Hospital,Fourth Military Medical University of Chinese PLA.MATERIALS:This trial was done in the Laboratory of Gastrointestinal Surgery,Xijing Hospital,Fourth Military Medical University of Chinese PLA between September 2001 and April 2004.Eighty-three male SD rats were involved in this trial.METHODS: Forty-seven rats were randomly chosen to prepare diabetic rat models by penile-intravenous injection of 65 mg/kg streptozotocin.Thirty-six successful model rats were randomized into 3 groups,with 12 in each group:IR group,donor IPC(DIPC)group and recipient with two hindlims IPC(RIPC)group.Twelve of the remaining 36 normal rats served as control group,and the other 24 rats were used as donors.Laparotomy was conducted only in control group,and pancreas transplantation was conducted in the other 3 groups In DIPC group,the splenic vessels of donors were blocked for 5 minutes and reperfused for 5 minutes twice before obtaining pancreas from donor;In the RIPC group, blood flow of two hindlimbs of recipients was blocked for 5 minutes and reperfused for 5 minutes before reperfusing the pancreas of donor,and this procedure was repeated 3 times.IR group was untouched.MAIN OUTCOME MEASURES:① On the 5th day after operation,6 rats were randomly chosen from each group to detect small intestinal permeability[expressed with plasm fluorescent-isothiocyanate-dextran(FITC-dextran)concentration]and absorption function(expressed with plasm xylose concentration).② On the 5th day after operation.blood was taken from the left 6 rats in each group to detect serum tumor necrosis factor-α(TNF-α) and nitric oxide(NO)level as well as superoxide dismutase(SOD)and amylase activity.Ileal mucosal tissue was taken to detect wet weight of small intestinal mucosa,the height and width of microvilli,malonaldehyde(MDA)level and myeloperoxidase(MPO)activity.At the same time,mesenteric lymph node,liver and splenic tissue were taken to perform bacterial culture.Bacterial translocation was observed.RESULTS:After supplement,72 rats were involved in the result analysis.①Plasm FITC-dextran concentration of IR group were higher than that in control group,DIPC group and RIPC group,respectively(P＜0.01).②Plasm xylose concentration in the IR group was lower than that in the control group,DIPC group and RIPC group,respectively(P＜0.01).③Bacterial translocation rate in the IR group was higher than that in the control group,DIPC group and RIPC group,respectively(P＜0.01).④Small intestinal mucosal injury degree in the IR group was lower than that in the other 3 groups(P＜0.01).⑤Small intestinal MPO activity and MDA level in IR group were significantly higher than those in the other 3 groups(P＜0.01). Serum SOD activity and NO level were lower but amylase activity and TNF-α 1evel were higher in the IR group as compared with the other 3 groups(P＜0.01).CONCLUSION:IPC of two hindlimbs in both donor and recipient can protect small intestinal mucosal barrier and reduce bacterial translocation rate after pancreas transplantation in rats.

BACKGROUND: Pancreas transplantation alone (PTA) is an effective therapy for diabetic patients who do not occur chronic complications. It's important to establish the stable PTA animal models to investigate immunologic tolerance or ischemic/reperfusion injury.OBJECTIVE: To establish the model of pancreas transplantation alone (PTA) with enteric drainage in rat.DESIGN: Grouping and controlled animal experiment SETTING: Department of Gastrointestinal Surgery, Xijing Hospital,Fourth Military Medical University of Chinese PLA MATERIALS: Totally 90 SD male rats, with the body mass of 250-320 g,were chosen in this study. 58 rats were induced by intravenous administration of streptozotocin (STZ) at a dose of 65 mg/kg via penile vein and the rats whose fasting plasma glucose exceeded 19.4 mmol/L for more than 2weeks were selected, 22 rats was successful. Rats were randomly assigned to 2 groups: control group (10 healthy rats) and group PTA consisted of 22diabetic rats, which received PTA from 22 normal donors.METHODS: This experiment was conducted at the laboratory of Department of Gastrointestinal Surgery, Xijing Hospital, Fourth Military Medical University of Chinese PLA from January 1999 to July 2004. The blood vessels reconstruction of PTA were performed using end-to-side anastomosis between the donors' abdominal aorta segment (abdominal artery and splenic artery) and recipients' abdominal aorta, and end-to-end anastomosis between the donors' portal vein segment (splenic vein) and recipients'left renal vein (use a cuff). Pancreas exocrine drainage was made by pancreas intestine anastomosis (Roux-Y).MAIN OUTCOME MEASURES: Body mass, food intake, water intake and fasting blood glucose were monitored 2 days before operation and 1,3,7,14 and 30 days after operation, and the failure causes were analyzed.RESULTS: 22 rats in the model group and 10 rats in the normal control the vein of the rats , very severe diabetic symptoms appeared in 22 rats:Body mass, food intake, fasting blood glucose was increased than that of cipients operation was (32.2±12.7) minutes and (63.4±15.9) minutes respectively. And the mean time of warm and cold ischemic time was 0minute and (48.6±18.3) minutes, respectively. 11 of the 22 cases (50%)died or lost their function of the endocrine within 1 month in Group PTA.The main complications were secondary pancreatitis and pancreas leakage after transplantation (7 cases, 31.8%). All successful recipients' blood glucose lowed on the 1st and recovered to be normal on the 3nd after transplantation (P ＜ 0.01), and their food intake, water intake and urine volume decreased and became stable 14 days later.CONCLUSION: This method can be used to establish relative stable animal model. Successful PTAs may improve the pancreatic endocrine function of the diabetic rats.

Pancreas transplantation alone(PTA) which dose not popularize in clinical therapy the insulin-dependent diabete mellitus patients who have not occurred end-stage complications,which was resulted in not only the technical but also the sociological and ethnics reasons.There are some reasons to restrict the development of clinical PTA at present.In this article the sociological and ethnics reasons to circumscribe the development of clinical PTA were discussed.

ObjectiveTo investigate the effect of ischemic preconditioning (IPC) in the posterior limbs of rats of donor' pancreas graft, and analyze the correlations with adenosine. Methods18 steptozozin (STZ)-induced diabetic SD rats were randomly assigned to 3 groups: group I/R (n=6) received pancreas transplantation alone (PTA), Group IPC (n=6) received pancreas transplantation alone exposed IPC with 5 minutes ischemic and 5 minutes reperfusion induced by ligating donor's posterior limb three times before ablating donors, Group IPC+DPCPX received same treatment with Group IPC, but separately injected adenosine A1 receptor antagonist 8-cyclopentyl-1, 3-dipropylxanthine (DPCPX) before IPC. The blood glucose, TNF-α in serum, MDA and MPO in pancreatic tissue were monitored before and after reperfusion, and apoptotice cells were stained by TUNEL technique 2 h after reperfusion.ResultsThe blood glucose, TNF-α, apoptotice index (AI), MDA and MPO of group I/R were higher than that of Group IPC and Group IPC+DPCPX after reperfusion(P<0.01), but those of Group IPC+DPCPX were not markedly different with Group IPC(P>0.05). ConclusionIPC in the posterior limbs of rats can protect rat pancreas graft from I/R injury during PTA. Adenosine do not participate in the signal mechanism of this protection.

ObjectiveTo observe the effect of ischemic preconditioning (IPC) on apoptosis of transplanted pancreas cells in rats.Methods6 normal SD rats were assigned as control group. 18 steptozozin-induced diabetic SD rats were randomly divided into 3 groups: the I/R group (n= 6, received pancreas transplantation alone), DIPC group (n=6, received pancreas transplantation exposed IPC with 5 min ischemic and 5 min reperfusion twice) and RIPC group (n=6, received pancreas transplantation exposed IPC with 5 minutes ischemic and 5 minutes reperfusion induced by ligating donors' posterior limbs three times before anastomosing vessel). The blood glucose in serum, superoxide dismutase (SOD), myeloperoxidase (MPO), TUNEL cells in graft were monitored.ResultsAfter reperfusion, compared with the I/R group, the mean blood glucose levels, MPO levels and apoptotice index of graft reduced, the mean SOD levels of graft heightened in DIPC and RIPC groups significantly (all P<0.01).ConclusionIschemic preconditioning induced by graft and ligating donors' posterior limbs can reduce apopotosis of transplanted pancreas cells.

BACKGROUND: During pancreas transplantation, ischemia/reperfusion (I/R) injury can lead to many complications, which directly threaten the survival of the donor pancreas and the receptor itself, and the serious ones may result in the failure of transplantation. Ischemic preconditioning can protect the target organs in the following ischemia, which has become one of the hot spots in investigating organ transplantation.OBJECTIVE: To observe the early protective effect of ischemic preconditioning on the I/R injury of the grafted pancreas in the rat, and analyze its correlation with apoptosis.DESIGN: A randomized control animal experiment.SETTINGS: Department of Gastrointestinal Surgery, Department of Anesthesiology, Xijing Hospital of the Fourth Military Medical University of Chinese PLA.MATERIALS: Seventy male SD rats of 3-6 months, weighing 250-320 g, were used.METHODS: The experiments were conducted in the laboratory of Department of Gastrointestinal Surgery between September 2001 and April 2004.Six normal rats were taken as the control group, and 24 successful diabetic models were divided into I/R group and 1, 2 and 3-time ischemic preconditioning groups (n=18) according to the method of random number table,with 6 rats in each. The rats in the latter three groups were treated with 5-minute ischemia and 5-minute reperfusion for once, twice and three times respectively, all the rats underwent the pancreas transplantation. Twentyfour SD rats served as donors.MAIN OUTCOME MEASURES: ① Blood glucose before and after reperfusion in each group; Serum contents of tumor necrosis factor alpha (TNF-α) and nitric oxide; Activities of superoxide dismutase (SOD) and myeloperoxidase (MPD) and content of malondialdehyde (MDA) in the grafted pancreatic tissue; ② Apoptosis in the grafted pancreatic tissue observed by means of in situ end-labeling; Expressions of Bax and Bcl-2 proteins in the grafted pancreatic tissue with the method of Western blotting.RESULTS: ① Changes of blood glucose before and after reperfusion: The levels of blood glucose were decreased as compared with those before reperfusion in the I/R group and ischemic preconditioning groups. It was significantly lower in the 2-time ischemic preconditioning group than in the I/R group, 1 and 3-time ischemic preconditioning groups (P ＜ 0.05). ②Serum content of TNF-α at 2 hours after reperfusion: It was lower in the ischemic preconditioning groups than in the I/R group; It was lower in the 2-time ischemic preconditioning group than in the 1 and 3-time ischemic preconditioning groups (P ＜ 0.05). ③ Serum content of nitric oxide after reperfusion: It was higher in the ischemic preconditioning groups than in the I/R group; It was higher in the 2-time ischemic preconditioning group than in the 1 and 3-time ischemic preconditioning groups (P ＜ 0.05). ④SOD activity in the grafted pancreatic tissue after perfusion: It was higher in the ischemic preconditioning groups than in the I/R group; It was higher in the 2-time ischemic preconditioning group than in the 1 and 3-time ischemic preconditioning groups (P ＜ 0.05). ⑤ MAD content and MPD activity in the grafted pancreatic tissue after perfusion: Those were lower in the ischemic preconditioning groups than in the I/R group, also lower in the 2-time ischemic preconditioning group than in the 1 and 3-time ischemic preconditioning groups. ⑥ Apoptosis in the grafted pancreatic tissue: The apoptosis index after perfusion was lower in the ischemic preconditioning groups than in the I/R group; It was significantly lower in the 2-time ischemic preconditioning group than in the 1 and 3-time ischemic preconditioning groups (P ＜ 0.05). ⑦ Expressions of Bax and Bcl-2proteins in the grafted pancreatic tissue: There was high expression of Bax protein and low expression of Bcl-2 protein in the grafted pancreatic tissue after perfusion in the I/R group; Low expression of Bax protein and high expression of Bcl-2 protein in the grafted pancreatic tissue after perfusion were observed in the ischemic preconditioning groups; In the 2-time ischemic preconditioning group, the expression of Bcl-2 protein was the highest but that of Bax protein was the lowest.CONCLUSION: Ischemic preconditioning can protect the grafted pancreas from I/R injury at early pancreas transplantation, which maybe correlated with the elevation of SOD activity, increase of the synthesis of endogenous nitric oxide, down-regulation of TNF-α and the alleviations of the adhesion and aggregation of polymorphonuclear leukocytes. Ischemic preconditioning can reduce the apoptosis of the grafted pancreas, and the the possible mechanism may be correlated with the alleviations of the adhesion and aggregation of polymorphonuclear leukocytes, reduce of oxygen-derived free radicals, up-regulation of Bcl-2 protein and the down-regulation of Bax protein. 5-mintue ischemia and 5-minute reperfusion for twice is the best way to induce ischemic preconditioning in rat pancreas transplantation.

Objective: To investigate the effect of interleukin-22 (IL-22) on the activation and proliferation of hepatic stellate cells (HSCs) induced by acetaldehyde, as well as the role of the antioxidant axis Nrf2-keap1-ARE. Methods: Hepatic stellate cell-T6 (HSC-T6) cells were cultured in vitro, and after 24 and 48 hours of acetaldehyde stimulation at various concentrations (25, 50, 100, 200, and 400 μmol/L), MTT assay was used to measure cell proliferation rate to screen out the optimal conditions for model establishment. HSC-T6 cells were treated first with the optimal concentration of acetaldehyde (200 μmol/L) for 24 hours and then with different concentrations of IL-22 (10, 20, and 50 ng/ml) for 24 hours. MTT assay was used to measure cell proliferation, Western blot and cell immunohistochemistry were used to measure the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and α-smooth muscle actin (α-SMA), and spectrophotometry was used to measure the changes in the content of malondialdehyde (MDA) and reduced glutathione (GSH) in culture supernatant. SPSS 17.0 was used for statistical analysis and data were expressed as mean±SD. P < 0.05 was considered statistically significant. A one-way analysis of variance was used for comparison of means between any two groups. Results: HSCs had significantly enhanced proliferation and activation after being treated with acetaldehyde, especially at 200 μmol/L for 48 hours. After the intervention with gradient concentrations of IL-22, the proliferation and activation of HSCs were inhibited in a dose-dependent manner, and the proliferation and migration rates in the 10, 20, and 50 ng/ml IL-22 groups were 14%, 25%, and 35%, respectively (all P < 0.05). The results of Western blot and immunohistochemistry showed that there was no significant difference in the expression of Nrf2 total protein in HSCs between groups, while there was extremely low expression of Nrf2 nucleoprotein in the blank control group. There was increased expression of Nrf2 nucleoprotein after acetaldehyde stimulation (compared with the blank control group, P < 0.05), and after the intervention with gradient concentrations of IL-22, the expression of Nrf2 nucleoprotein was further increased (all P < 0.05). The results of spectrophotometry showed that compared with the blank control group, the model group had increased levels of MDA and GSH in culture supernatant after acetaldehyde stimulation; after the intervention with gradient concentrations of IL-22, there was a significant reduction in the MDA level and a significant increase in the GSH level in a dose-dependent manner (all P < 0.05). Conclusion The activation and proliferation of HSCs induced by acetaldehyde helps with the successful establishment of an in vitro model of alcoholic liver fibrosis. IL-22 effectively inhibits the activation and proliferation of HSCs induced by acetaldehyde, and its mechanism may be related to promoting Nrf2 nuclear translocation in HSCs and expression of the downstream target gene GSH and increasing the activity of the antioxidant axis Nrf2-keap1-ARE.

Objective To investigate the value of three-dimensional speckle tracking imaging(3D-STI) in assessment of left ventricular regional systolic function in acute myocardial infarction model of dogs.Methods Left anterior descending (LAD) coronary artery was ligated in 24 adult beagle dogs.Longitudinal strain(LS) of every left ventricular segment before ligation and 180 min after ligation,and the time for acquisition and analysis were obtained using two-dimensional speckle tracking imaging (2D-STI) and 3D-STI.After the experiment,left ventricular myocardium were determined by TTC staining and divided into two groups:infarcted myocardial segments and non-infarcted myocardial segments.Results Experimental model of acute myocardial infarction was established successfully in 20 dogs.After ligation,both LS of 2D-STI and 3D-STI in infarcted myocardial segments were significantly lower (P ＜ 0.01).The time for acquisition and analysis of the images by 3D-STI were significantly shorter than that by 2D-STI (P ＜0.01).The area under the ROC curve of 3D-STI was larger than that of 2D-STI.A cut-off value of-12.56％ for LS of the 3D-STI had 77.9％ sensitivity and 88.2％ specificity in predicting acute myocardial infarction.Conclusions 3D-STI is a convenient,accurate method for evaluating left ventricular regional systolic function.