Effects Of Heparin On Hepatic Regeneration After 50% Partial Liver Transplantation In Rats

【Objective】Unfractionated heparin,low molecular weight heparin were regularly utilized to prevent or treat hepatic artery thrombosis during early postoperative in clinical living donor liver transplantation.However,there is few literatures on effects of heparin on hepatic regeneration after 50%partial liver transplantation in rats.This study was designed to investigate effects of unfractionated heparin on liver regeneration in rats after reduced-size liver transplantation.【Methods】Try to set up stable Orthotopic Liver Transplantation model and serial reduced-size liver transplantation(60%,50%,30%) model in rats using two-cuff technique.To master the surgical skills of Liver transplantation in rats and reduce the complications.On this basis to select the same season,the same batch healthy male wistar rats 120(Beijing Academy of Military Medical Sciences),weighting 230-280g, were randomly divided into four groups(random number table method):A,B two sets were 42 rats each group,C,D were 18 in each group,the donors and receptors were randomly selected.50%reduced-size orthotopic liver transplantation was performed in A,B groups using two-cuff technique.The rats in A,B groups were given normal saline(4ml/kg) and unfractionated heparin(1000U/kg) respectively. 50%hepatectomy was performed in C group given saline after hepatectomy(4ml/kg). D group was sham-operated group,given normal saline(dose with the C group) postoperative.All of the drugs were administrated subcutaneous 20h postoperative. Rats in each group after 3 days,5 days,7 days were randomly sacrificed 6.Blood samples(5ml) were obtained from inferior vena cava of rat when Rats were sacrificed after anesthesia.Get serum after standing and centrifugalization,then put serum in the refrigerator(-80℃),and detect hepatocyte growth factor(HGF) by enzyme-linked immunosorbent assay(ELISA);check transplanted liver,Record rat weight,liver wet weight,the calculation of liver regeneration rate,make single-cell suspension from part of a liver by mechanical method;Detect Liver cell proliferation index by flow cytometer,part of a liver cryopreserved -80℃,and the remaining part of liver was fixed by formaldehyde,make conventional HE staining and immunohistochemical staining(liver cells proliferating nuclear antigen PCNA),Record PCNA expression. Results were analyzed using SPSS 13.0 statistical analysis software,observe compa rison of various indicators of group differences in liver regeneration(independent samples t test),and compare each group differences in the levels of HGF.【Results】Complete one hundred cases of orthotopic liver transplantation in rat and a series of reduced-size liver transplantation(60%,50%,30%) before formal experiment,and set up a stable model of rat liver transplantation.Rats of group A and B were performed 50%reduced-size liver transplantation,the time for donor operation was(30±3)min and(31±4)min respectively,the recipient operation time was(54±7)min and(58±5)min,respectively,the anhepatic time was(15±2)min and(16±2)min,cold ischemia time was(56±5)min,(54±7)min respectively,the 48 hours survival rate was 95.2%(20/21)and 100%(21/21),no rats died of infection and bleeding.Two rats of group A were excluded due to biliary complications of postoperative,the incidence of biliary complications was 10%(2/20).Three cases of group B were excluded due to biliary complications of postoperative,the incidence of biliary complications was 14.3%(3/21).The rats of group C(18 cases ) were completed 50%hepatectomy,time for the operation was(14±2)min.There was no case of death and complications of group C and D.Liver regeneration rate of groups A,B,C on the third,fifth,seventh day after surgery was(33.5±6.6)%,(43.7±5.4)%,(45.6±6.3)%,(40.1±2.8)%,(71.9±17.8)%,(56.9±5.4)%,(86.6±11.9) %,(111±19.2)%,(98.4±4.6)%,respectively.There were significant differences(P = 0.015 <0.05,P= 0.002 <0.01,P= 0.023 <0.05) between A and B on the third,fifth, seventh day after surgery.There were significant differences(P=0.012<0.05,P= 0.011<0.05,P=0.031<0.05) between A and C on the third,fifth,seventh day after surgery.Serum HGF levels of groups A,B,C on the third,fifth,seventh day after surgery were(263.3±101.1)pg/ml,(420.7±56.8) pg/ml,(365.3±6.4)pg/ml, (312.9±10.1)pg/ml,(440.9±23.4)pg/ml,(384.4±32.2)pg/ml,(242.4±22.1) pg/ml,(409.9±23.0)pg/ml,(376.3±22.3)pg/ml.There were significant differences on serum HGF levels on the third,fifth,seventh day after surgery(P = 0.008 <0.01,P = 0.000 <0.01,P = 0.000 <0.01) between A and B.There were significant differences on serum HGF levels on the third,fifth,seventh day after surgery(P=0.016<0.05,P=0.038<0.05,P=0.001 <0.01) between A and C. Hepatocyte PI index of groups A,B,C on the third,fifth,seventh day after surgery were as follows:(30.4±4)%,(48.1±8)%,(38.4±2)%,(29.3±3)%,(43.5±8) %,(36.9±2)%,(30.3±3)%,(47.3±8)%,(31.6±3)%,there were significant differences on hepatocyte PI index on the third,fifth,seventh day after surgery(P = 0.001 <0.01,P = 0.002 <0.01,P = 0.02 <0.05)between A and B,and significant differences(P=0.025<0.05,P=0.018<0.05) on the third,fifth day between A and C,but have no difference on the seventh day between A and C(P=0.86>0.05).There were differences on PCNA expression between A and B on the third,fifth day,but no significant difference on PCNA expression on the seventh day.There were differences on PCNA expression between A and C on the fifth day,but no significant difference on PCNA expression on the third,seventh day.【Conclusion】unfractionated heparin(1000IU/kg) given subcutaneous significantly promote hepatic regeneration after 50%reduced-size liver transplantation in rats;the liver regeneration of 50%reduced-size liver transplantation was delayed comparing to hepatectomy.In this experiment,it may be one of the mechanisms beneficial to liver regeneration that heparin elevates the plasma production of hepatocyte growth factor.