Objective To investigate the joint effects of selective digestive decontamination (SDD) and glutamine (Gln) on preventing intestinal bacterial translocation of orthotopic piggyback liver transplantation and to observe the incidence of postoperative pneumonia in rabbit. Methods Thirty rabbits received orthotopic piggyback liver transplantation and were randomly divided into three groups (SDD group, SDD+Gln group and control group). Mixed emulsion of tobramycin, polymyxin E and nystatin were given to the rabbits in SDD group. Same dosage of the above components plus Gln were given to the rabbits in SDD+Gln group. Samples of portal vein blood, ileum tissue and lung tissue were obtained in each group at different phases during and after operation, the pathological changes of ileum tissue, the bacterial translocation in blood of portal vein and the incidence of postoperative pneumonia were detected. Results The mixing section area of intestinal blood capillaries in SDD+Gln group was smaller compared with control group (P＜0.05， P＜0.01) and SDD group (P＜0.05) while the portal vein was obstructed for 15, 30 and 45 min, and after the operation, respectively. The length of ileum villus in SDD+Gln group was longer than that in control group (P＜0.05) and in SDD group (P＜0.05) before the portal vein was obstructed, but the length of ileum villus in control group gradually became longer and eventually exceeded that in SDD+Gln group at the time of 45 min after the portal vein was obstructed (P＜0.05). After the operation, the length of ileum villus in SDD+Gln group was significantly longer than that in SDD group (P＜0.05) and control group (P＜0.01). At the time of 45 min after the obstruction of portal vein and 30 hours after operation, the positive rate of cultured bacterial in the blood of portal vein in SDD+Gln group was significantly lower than that in control group (P＜0.05, P＜0.01). The incidence of postoperative pneumonia in SDD+Gln group and SDD group were significantly lower than that in control group (P＜0.05,P＜0.01). Conclusion Gln could nourish intestinal epithelium of mucous membrane.When combined with SDD, it could decreased the incidence of intestinal bacterial translocation occurred during the obstruction of portal vein and after operation, so as to decrease the incidence of postoperative pneumonia.
Objective To study the mechanism of immune hyporesponsiveness of allograft rejection induced by transfusion nonpufsed allopeptide syngeneic immature dendritic cell (imDC) generated from recipient bone marrow progenitors and to explore a possible strategy for liver allograft protection in clinic. Methods Forty experimental rats were randomly divided into 4 group: control group, cyclosporine A (CsA) group, mature DC (mDC) group and imDC group. In control group, Wistar rats only received liver transplantation. In CsA group, Wistar rats underwent liver transplantation plus CsA treatment 〔10 mg/（kg·d)〕. In mDC group, recipient-derived mDC 1×106 were infused intravenously through the penile vein to Wistar rats. In imDC group, ImDC with the dose of 1×106 were injected into Wistar rats via the dorsum vein of penile. In each group, five recipients were killed on the 10th day after transplantation, the other five recipients were left to observe survival time. The levels of ALT, AST, TBIL, IL-2, IFN-γ, IL-4 and IL-10 were detected. The acute rejection and the expression of FasL/Fas in the grafts were detected by HE and immunohistochemical staining. Western blot was used to detect Scurfin protein expression of CD4+ CD25+ T cells. Results The median survival time of the liver allografts in CsA group and imDC group were significantly longer than that in control group and mDC group ( P ＜ 0.05). The levels of ALT and TBIL in control group and mDC group were significantly higher than those in CsA group and imDC group ( P ＜ 0.05). Compared with CsA group and imDC group, the levels of IL-2 and IFN-γ were higher but the levels of IL-4 and IL-10 were lower in control group and mDC group ( P ＜ 0.01). Slightly or no rejection reaction was found in CsA group and imDC group ( P ＜ 0.05). The Scurfin protein expressions of CD4+ CD25+ T cells of imDC group were significantly higher than those of other three groups. Conclusion Application of nonpufsed allopeptide syngeneic recipient-derived imDC is an effective way to induce immune hyporesponsiveness by blocking indirect recognition in rat liver transplantation model. Survival span is significantly prolonged by its protective effect. The mechanism of immune hyporesponsiveness induced by imDC transfusion might be involved in some aspects: T cell apoptosis, immune deviation of Thl/Th2 cytokine net and inhibition of T lymphocytes responsiveness by regulatory T cells.
Objective To explore improvement of orthotopic liver transplantation model in rhesus monkey. Methods Healthy rhesus monkeys were chosen to perform orthotopic liver transplantation for 10 cases. The model was established by drawing on a variety of animal model methods, and the portal vein cuff method was used to establish stable model of orthotopic liver transplantation in rhesus monkeys. Results Ten orthotopic liver transplantation models in rhesus were performed, and the achievement ratio of operation was 10/10. The time of donor hepatectomy and donor preparation was (20±5) min and (30±7) min, respectively. The operation time of recipient and anhepatic phase were (180±35) min and (17±4) min, respectively. After 24 h of operation 9 cases survived, one case died of intra-abdominal hemorrhage after 9 h of operation. After 72 h of operation 8 cases survived, and one case died of upper gastrointestinal bleeding after 38 h of operation. After one week of operation 5 cases survived, and 3 cases died of rejection after 9, 11, and 11 d of operation, respectively. The longest survival time was 32 d, but all of them also died of rejection. No portal vein thrombosis and biliary complications were found in all recipients.Conclusion The improved rhesus monkey model of orthotopic liver transplantation is easy to perform with high achievement ratio of operation. It is an ideal animal model for pre-clinical studies of liver transplantation.
Objective To explore the methods of hepatic artery reconst ruction with iliac arterial interpositiongraf t in orthotopic liver t ransplantation (OL T) and influential factor of relevant complications postoperatively.Methods Analyzed ret rospectively 8 OL T , the hepatic artery reconst ruction with arterial inflow based on recipientinf rarenal aorta using donor iliac artery graf t tunneled through the t ransverse mesocolon and pancreas. Results Thetime required for hepatic artery reconst ruction with iliac arterial interposition graf t was 52 - 126 minutes. Amongthe 8 patient s , 2 patient s developed postoperative bililary t ract complications , 1 with biliary fistula , 1 with int rahepatic biloma , the others were recovered smoothly and liver function returned to normal about one week af ter livert ransplantation. No complications of hepatic artery were observed. Conclusion Iliac arterial interpositional graft is aneffective and reliable method of revascularization in liver transplantation when the use of hepatic artery is not possible.
Objective To observe the dynamic histopathologic changes of acute rejection in rat orthotopic liver transplantation (OLT) model after tacrolimus discontinued and provide some prediction and evaluation data for clinical acute rejection after liver transplantation. Methods Kamada two-cuff technique was used to establish 60 rat OLT model， and male DA rats, male Lewis rats were used as donors and recipients respectively. Therapeutic amount of tacrolimus (0.05 mg/kg, twice per day, continued for 8 d, 1 d before operation and 7 d after operation, intragastric administrated) was administrated to recipients, then continuously half dose was decreased every day beginning from day 8 after operation and tacrolimus administration was stopped on day 13. Liver tissues were collected on day 7, 14, 21, and 28 after liver transplantation. Histopathologic changes and rejection activity index (RAI) of liver tissues were observed, survival time of recipients was calculated. Results Owing to protection effects of tacrolimus, liver tissues displayed no significant histopathologic changes of acute rejection in 7 d after OLT, while typical acute rejection histopathologic changes began to be observed on day 14 after OLT due to tacrolimus discontinuation. On day 14, 21, and 28, RAI were 3.7±0.9, 6.3±0.9, and 8.1±0.7 respectively. Survival time of recipients was (20.85±0.71) d with a median of 21 d. Conclusion Acute rejection could be induced in rat OLT model after tacrolimus discontinuation, and data collected from this model shows some extent of predictive value and assessment value for clinical liver acute rejection.
ObjectiveTo clone full-length cDNA of rat galectin-9 and construct recombinant adenovirus granule containing rat galectin-9 gene. MethodsThe galectin-9 gene was amplified by RT-PCR from rat liver tissue and inserted orientationally into plasmid pDC316-GFP digested by restriction endonucleases NotⅠ and HindⅢ. The recombinant pDC316-GFP-galectin-9 shuttle plasmid was identified by PCR, restriction endonuclease digestion and sequencing, and then co-transfected with rescue plasmid pBHGlox△E1.3Cre into HEK-293 cells by liposome reagent. Recombinant adenovirus vector containing rat galectin-9 gene (Ad5-galectin-9) was generated by sitespecific recombination and confirmed by PCR, and then Ad5-galectin-9 was propagated in HEK-293 cells and purified. The infectious titer of viral stock was determined by TCID50 assay. ResultsConstruction of pDC316-GFP-galectin-9 shuttle plasmid was confirmed to be correct by PCR, restriction endonuclease digestion and sequencing. Construction of recombinant adenovirus Ad5-galectin-9 was confirmed to be correct by PCR. The infective titer of Ad5-galectin-9 was 1.4×109 U/ml. ConclusionRecombinant adenovirus vector containing rat galectin-9 gene (Ad5-galectin-9) is successfully constructed, which provides the foundation of further research on the function of galectin-9 gene.
ObjectiveTo explore perioperative management model of ABO-incompatible liver transplantation. MethodsThe clinical data of ABO-incompatible caderveric liver transplantions without urgency performed in our center from July 2006 to May 2010 were analyzed retrospectively. Four patients had received an ABO-incompatible graft: AB to O in three, AB to A in one. All the cases were diagnosed as end-stage liver disese, one of them was primary hepatocellular carcinoma. ResultsFour survived to now （11 to 19 months） without severe infections and acute rejections. Two experienced coagulative disturbance and one of them had a second exploration. One developed acute renal failure and recovered with help under continuous veno-venous hemofiltration. All the cases were given 20 mg basiliximab two hours before revascularization and on day 4 after operation respectively. Splenectomy was performed in three, intravenous immunoglobulin was given in all more than seven days. Isohemagglutinin titers were basically stable and not relevant to the clinical manifestations. Antibiotic prophylaxis and immunosuppression protocol was same as the ABO compatible transplants except a 3-month-delay for steroid withdrawal. ConclusionABO-incompatible liver transplantation could be performed with appropriate perioperative management, such as basiliximab induction, splenectomy, intravenous immunoglobulin administration, and routine immunosuppression.
Objective To explore the methods of hepatic artery reconstruction in orthotopic liver transplantation （LT） and prevention of relevant complications postoperatively. Methods A retrospective analysis was made for 31 cases orthotopic LT. Results The variations of hepatic arteries, which did not exist in the recipients, were found in 2 living donors. In 1 case, the accessory left hepatic artery arose from the left gastric artery. The ends of accessory left hepatic artery and left hepatic artery were made into one end through angioplasty on the back table. An interposition graft of donor great saphenous vein was used in the arterial reconstruction. In the other cases, the accessory right hepatic artery originating from gastroduodenal artery and the right hepatic artery were anastomosed to the branches of the hepatic artery of the recipient separately. In 1 patient receiving dual graft LT, the arteries of the grafts were nastomosed to the branches of the hepatic artery of the recipient separately. The diameters of hepatic arteries were less than 3 mm in 6 cases and more than 5 mm in 8 cases, the others were 3 to 5 mm. Donor iliac arterial graft was used for interposition between graft hepatic artery and recipient abdominal aorta in 1 case. Microsurgical vascular techniques was utilized in the reconstruction of hepatic artery. The time for an arterial reconstruction was 23-70 min 〔（31.46±9.07） min〕. The patients were followed up for 2-7 months. Hepatic artery stenosis was detected in 1 case on 32 d after LT, and no other arterial complications were found. Conclusion To attach importance to factors contributing to hepatic artery complications, the microsurgical technique applied in the reconstruction of the hepatic artery and appropriate anticoagulation can help to prevent the hepatic artery complications in LT.