| Miniature pig was ideal experimental animal for its similarity with human anatomy, physiology, biochemistry and disease mechanism. The experimental liver injury animal models of miniature pigs were the urgent needed problem to resolve in veterinary medicine and comparative medicine. This topic proceed a pioneering research in the building of liver injury animal models in miniature pigs by laparoscopic minimally invasive surgical technique, and explore the operation method of laparoscopic liver resection and ischemia-reperfusion injury model, and monitoring the changes of the liver environment of the injury. We also compared laparoscopic and open model at the same time, and further to evaluate scientific and minimally invasive advantage of the model, this also lay the foundation for the laparoscopic technique in building miniature pig experimental models.This experiment selects 40 miniature pigs as experimental animals, randomly selected five pigs for laparoscopic abdominal cavity surgical exploration. Then the remaining 35 pigs were randomly divided into 5 groups, each group of seven, respectively, the laparoscopic majority liver resection model group and its control group, the laparoscopic liver ischemia combined resection model group, laparoscopic liver resection group and the control group. By pneumoperitoneum, surgical access, separation of ligaments, the anatomy of the first porta hepatis, throughout the liver parenchyma and hemostasis, right hepatic ischemia, cutting off the liver parenchyma, washing of the abdominal cavity, resection of liver taking out, the liver injury models were established.Intraoperative of vital signs, circulation system, respiratory system and body temperature were entire continuous monitored and record the intraoperative and postoperative complications, operation and postoperative recovery, after 30 d laparoscopic quadratic probe surgical site adhesion. Between groups in preoperative, 4h, 1d, 3d, 7d and 14 d, 30 d take venous blood, detection routine blood and serum total protein, aspertate aminotransferase, r-glutamine transferase, total bilirubin, urea nitrogen, creatinine, and plasma coagulation function; In the preoperative and postoperative immediate, postoperative 1d, 3d, 7d after operation and postoperative 1 month after surgery, laparoscopic liver biopsy were taken, paraffin section and HE staining to observe the histopathological changes, at the same time for different model of liver tissue was detected PCNA and cyclin D1, tumor necrosis factor-alpha and interleukin-6 expression and organization of malondialdehyde content, superoxide dismutase, catalase activity and Caspase-3 activity, statistics and contrast the results of laparoscopy and laparotomy and serum cortisol, the changes of c-reactive protein.Laparoscopic liver partial operative field exploration and anatomical observation found that small liver of pig anatomical location, shape structure, ligament distribution, pipeline system and lobe division and human approximation, its unique anatomical structure to facilitate the liver model.This subject successfully completed the miniature pig most laparoscopic liver resection liver ischemia model and miniature pig laparoscopic combined resection model, intraoperative and postoperative physiological have degree of volatility, but neither is beyond the scope of miniature pigs safety tolerance; Relative to laparotomy, laparoscopic surgery time is a bit long, but the small incision, less blood loss, postoperative abdominal adhesions, and all the miniature pigs have a rapid recovery, postoperative mental status of 1d and appetite to eat a small amount of liquid feed, postoperative 7-9 d the abdominal incision for the first phase of healing.After model establishment, the mainly environmental changes of majority liver resection model are: the postoperative serum AST extremely significant difference compared with the preoperative and the control group(P<0.01) for 1d and 3d after operation, the significant difference(0.01<P<0.05) 7d after operation. Pathology observation are liver cell swelling, degeneration and inflammatory cell infiltration, hepatic tissue damage is obvious7 d after surgery; Postoperative 1d hepatocellular PCNA expression level in peak, then gradually reduced. Amount of liver tissue Cyclin D1 gene expression in liver most early expression increased significantly after resection, postoperative 1d increased significantly(P<0.01), which continues to postoperative 3d(0.01<P<0.05).The postoperative liver tissue TNF alpha reduce after rising first, 1d lifts quickly after surgery to the peak, the extremely significant difference compared with the control group(P<0.01), 3d, 7d after gradually reduce, significant difference(0.01<P<0.05); Small variations in IL-6, 1d and 3d significant difference compared with control subjects(0.01< P <0.05).Liver environment changes of liver ischemia combined resection model are: compared with the control group 1d and 3d postoperative serum AST extremely significant difference(P<0.01), postoperative7 d significant difference(0.01<P<0.05), and other monitoring point in time no significant change(P>0.05).Postoperative 1 d model group remaining liver tissue has a wide range of liver cell degeneration and mild atrophy, occasional sinus gap passive congestion, hepatic lobule with local focal hepatocyte necrosis or patchy necrosis, interstitial inflammatory cell infiltration; Postoperative visual local lobular necrosis area within 7 d liver cell proliferation, local portal area fibrous connective tissue hyperplasia, etc. Compared with control group, in the immediate postoperative malondialdehyde rise significant difference(P <0.01), intraoperative and postoperative 1h,1d liver tissue malondialdehyde changes significant difference(0.01<P<0.05), and other monitoring point in time no significant change(P>0.05); In the immediate postoperative superoxide dismutase changes extremely significant difference(P<0.01), postoperative 1d changes significant difference(0.01<P<0.05), and other monitoring point in time no significant change(P>0.05); In the immediate postoperative catalase extremely significant difference(P<0.01), intraoperative 1 h,1d after significant difference(0.01<P<0.05), and other monitoring point have no significant change(P>0.05). Compared with the control group, Caspase-3 activity in liver tissue changes difference was significant(P<0.01) in postoperative 1d, intraoperative and postoperative immediately 3d change significant difference(0.01<P<0.05), no significant change in other time points(P >0.05). Through the experimental results, the following conclusions were drawn:(1) The use of laparoscopic minimally invasive surgery technology can successfully built the majority liver resection model. ansd liver ischemia combined resection model in miniature pigs(2) Through the comprehensive monitoring of perioperative vital signs, circulation system, respiratory system, liver and kidney function, blood coagulation function, the physiological indexes are maintained in a safe range, proved that the technology and operation method is safe and feasible.(3) Compared with traditional laparotomy, laparoscopic surgery have small incision, less blood loss, postoperative adhesion is light, and the stress response to animals is small, with rapid postoperative recovery and minimally invasive advantage.(4) After model establishment, through the test and evaluation of liver tissue and serum, we clear the changing rule of liver function, pathology, and liver cell proliferation, cytokines and growth factors, oxidative stress and apoptosis after injury. |
