| The difficulty of small bowel transplantation skills is well known, as intestinal lymphoid tissue is rich and a large number of microorganisms is contained in the intestine. Due to these reasons, small bowel transplantation is considered taboo to scientists for a long time. However, with the fast development of surgical techniques in recent decades, small bowel transplantation has already received a lot of success. The Starzl Transplantation Institute, University of Pittsburgh is a world famous small bowel transplantation center; 10-year’s graft survival rate has reached to 61% while 15-year’s survival rate reaches to 50%.With the improvement of surgical techniques, immunosuppressive drugs and infection control, small bowel transplantation will certainly has a broader future. This is because there is still a great demand for small bowel transplantation in the world. Many patients can not adapt total parenteral nutrition (TPN), yet they can ultimately get rid of it. In this case, small bowel transplant is a last resort to save lives, and its development is high of importance.Healthy human gut has a Gram-negative Proteobacteria and Bacteroidetes, and Gram-positive Firmicutes. Clostridium and Lactobacillus are both the main categories of intestinal bacteria. Gut flora has played an important role in the maintenance of normal intestinal flora, regulation of host immune and organ metabolism, impaction of host healthy growth and physiological function. Once the balance of intestinal flora has been destroyed, harmful bacterias would translocate into the blood and worse the disease. Small bowel transplantation has its specificity, as the gut flora will has a tremendous change during perioperative period. What is the law of gut bacteria change? How to early detect and treat flora disturbance, thus to prevent harmful gut bacterial translocation is the focus and difficulty in recent research.Small bowel transplantation exists an imbalance of intestinal flora, and intestinal mucosal barrier injury. These can be alleviated by using of antibiotics and probiotics (including probiotics, prebiotics, etc.). Probiotics can not only to antagonize pathogene and nutrition gut, but also to improve the function of macrophages, NK cells, and stimulate S-IgA secretion. Some animal experiments and clinical studies reported that application of probiotics can reduce the incidence of infection after transplantation. Therefore, using probiotics to prevent diseases and infections is a leading-edge research in this area. The main subject of our study based on PCR-DGGE technique to detect intestinal flora and intestinal epithelial barrier changes, and observate the variation law behind. We also study the impact of antibiotics and probiotics using perioperatively on gut flora change, to find new ideas and new strategies to prevent the infections of small bowel transplantation.Chapter 1 Characteristic changes of intestinal microflora and epithelial in acute rejection of small bowel transplantationObjective:Bacterial translocation is mainly dependent on the changes of intestinal flora in acute rejection of small bowel transplantation. The main part of this section is to reveal the feature of ileum flora in acute rejection of small bowel transplantation.Methods:A randomized into three groups:1) Sham group (n= 6):BN rats are used. We open the abdominal cavity and then suture;2) Syngeneic transplantation group (n= 6):BN rats are used both for donor and acceptor;3) Allogeneic transplantation group (n= 6):Lewis rats are used as donor, and BN rats for receptor.Using denaturing gradient gel electrophoresis (DGGE) studies to reveal ileum flora changes in Day 7 after modeling. Pathological examination using transmission electron microscopy and general morphology of the intestinal mucosa and tight junction structural changes using microscope are conducted. Immunohistochemical study of tight junction protein Claudin-1 is also administered.Results:DGGE analysis showed ileal flora changed significantly during acute rejection of rat small bowel transplantation model. Allogeneic transplantation group and the sham group was significantly different. The flora is characterized that E. coli and other harmful bacteria increased significantly in ileum, while Lactobacillus and other beneficial bacteria was decreased. Allogeneic group had an impaired mucosal barrier function, a damage of intestinal integrity, and a wider tightly coupled gap compared with the sham group. The difference was significantly.Conclusion:During acute rejection of small bowel transplantation, intestinal flora was apparently disordered, and intestinal barrier function was also impaired.Chapter 2 Perioperative drugs use in changing gut flora and regulating epithelial in acute rejection of small bowel transplantationObjective:In clinical, perioperative use of antibiotics is essential during small bowel transplantation. Besides, more and more probiotics are emerging used. These interventions on gut flora and intestinal barrier function will inevitably have a significant affect. However, it is still not systematically widely reported and studied now. Our study focused on two representative pharmaceutical, gentamicin and Bifico, to study whether and how to intervene gut flora change and bacterial translocation.Methods:A randomized into three groups, each rat received drugs intragastricly 14 days around the small bowel transplantation.1) Saline group (group C, allogeneic+NS group, n= 6):allogeneic small bowel transplantation (Lewis-BN)+2ml saline gavage;2) Antibiotic group (group D, allogeneic+Gen group, n= 6):allogeneic small bowel transplantation (Lewis-BN)+2ml gentamicin gavage;3) Probiotic group (group E, allogeneic+Bifid group, n= 6):allogeneic small bowel transplantation (Lewis-BN)+2ml Live Combined Bifidobacterium, Lactobacillus and Enterococcus Capsules, gavage.Use DGGE to study intestinal flora variation in rat ileum on Day 7 after modeling. Pathological examination using transmission electron microscopy and general morphology of the intestinal mucosa and tight junction structural changes using microscope are conducted. Immunohistochemical study of tight junction protein Claudin-1 is also administered.Results:DGGE analysis showed fecal flora diversity in antibiotic group were significantly different from those in saline group and probiotic group. In more detail, beneficial bacteria were reduced while pathogens were increased. Immunohistochemistry showed tight junction protein Claudin-1 in ileal epithelial in the probiotic group had been repaired; electron microscopy showed ileal epithelial tight junctions in the antibiotic group had a significantly widen gap.Conclusion:Perioperative use of antibiotics and probiotics during small bowel transplantation will lead a great change in ileal flora stucture. Antibiotics exacerbated the change and damage, while probiotics reduced these injuries.ConclusionAfter orthotopic small bowel transplantation model is well modeled, gut flora has an apparent disorder and structure change, which showed a significant increase in harmful bacteria, and reduce in most beneficial bacteria. The mucosal barrier function in allogeneic group is impaired, indicating those gut floras disorders are also exist. Flora disturbance and increase of epithelial tight junction’s gap can both lead to the occurance of gut bacteria translocations.Gentamicin and Bifico intervention have a great relationship to gut flora change. Fecal flora diversity in antibiotic group is significantly reduced while probiotics reduced this effect. Immunohistochemistry and electron microscopy showed that ileal epithelial tight junction protein and the gap in the probiotic group had both repaired. Above all, flora interventions have a significant impact on bacterial translocation in acute rejection of intestinal transplantation. |
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