| Objective:Wound healing is a huge reaction system involved in a large number of cells, cytokines, signaling proteins and related genes,and the differences within the different organizations environmental may cause different healing outcomes, and the unique physiological and anatomical features of bile duct caused a form of the healing outcome of scar, scar contracture will lead to the occurrence of bile duct stenosis, and then bile excretion or secretion occurs obstruction, which led to a series of clinical symptoms, eventually leading to biliary infection, biliary cirrhosis, liver failure and other outcomes.The aim of this issue is by establishing the healing model after bile duct injury in dogs, through the dynamic observation to during the scar formation after bile duct injury, the expression and location of Nuclear Factor Kappa B activation and its downstream apoptosis regulatory protein:cellular inhibitor of apoptosis protein-1,FLICE-like inhibitory protein and their downstream apoptotic factor caspase3,8 and the mechanism of bile duct wound healing and scar formation was expounded from the point of view of the the balance of proliferation and apoptosis, in order to provide ideas to find new ways for the clinical treatment of biliary stenosis. Methods:30 local health hybrid dogs, were randomly (number table) divided into experimental group (n=15) and control group (n=15), each further divided into 2months,3months,4months,5months,6months group.The models of wound healing after bile duct injury in dogs bying transverse to cut the whole floor of anterior wall and sides of common bile ducts using the electric knife, retaining hind wall.and then all underwent full thickness suture from mucosal to mucosa; the common bile ducts of the control group were only dissociated before that the abdomens were closed. After anastomotic bile duct tissue were drawn at the corresponding time points of each group after operations, immunohistochemistry SP method was using to detect and analysis the expression and localization of activated NF-κB,c-FLIP protein and caspase3,8,and in situ hybridization method was using to detect and analysis the expression and localization of cIAP-1 and c-FLIP mRNA expression. By image analysis system,the number of positive cells were quantitatively analysised both the groups.finally, statistically analysis and comparation were made.Results:1.The activation of NF-KB were significant at all the time points in the experimental group, Locating in the stromal cells, especially in fibroblasts.There were more significant difference between the experimental group(37.52±4.33～50.41±4.17)and control group(8.68±0.78～10.17±1.27)at the same phase (P<0.05),and the former was significantly stronger than the latter; There were significant differences within each time point of the experimental group in totality (P<0.05).Among them, the strongest group was 3months group(50.41±4.17%);there were no differences within 2 (P>0.05),3(P>0.05),4(P>0.05)months groupand were differences between 5 months and 2(P<0.05),3(P<0.05)months group;so is also between 6 months and 2(P<0.05),3(P<0.05),4(P<0.05)months group.2.There were all strong positive expression of cFLIPmRNA in the interstitial tissue of experimental group in all time points, mainly expressed in the cytoplasm of fibroblast and very little or almost no expression in the glandular tissue. Positive expression of cFLIPmRNA in the interstitial tissue(24.76±4.52～30.57±6.17)was significantly stronger than that of glands tissue(3.27±1.17～3.83±1.04) (P<0.05).There were no significant differences among each time point in either the interstitial tissue or glandular tissue (P>0.05).There were all weak expressions in the interstitial tissue(3.19±0.70～3.95±1.10)and glandular tissue(3.44±0.59～3.78±0.71)of the sham operation group at all time points and was no significant difference (P>0.05),no difference between each phase (P>0.05);The expression of the experimental group was significant (P<0.05) compared with sham operation group in the interstitial tissue at all time, while no that in the glandular tissue (P>0.05)3.There were all strong positive expression of c-FLIP protein in the anastomotic tissue of the experimental group at all time points(17.23±3.53～22.33±3.40),mainly expressed in the cytoplasm of fibroblastand very little or almost no expression in the glandular tissue. There were more significant difference between the experimental group and control group(3.11±1.01～3.41±0.69) at the same phase (P<0.05),and the former was significantly stronger than the latter; There were all weak expression of caspase8 in the anastomotic tissue(3.20±0.86～4.01±0.88)of the experimental group at all time points,significantly different compared with the control group(10.03±1.93～11.66±2.80)at the same phase(P<0.05;and there were no significant differences among all the time points(P>0.05).The expression of caspase8 in the experimental group showed poor. Most targeted to epithelial and stromal tissue is relatively less.The results of the correlation analysis:There was negative correlation in the expression between c-FLIP and caspase8 (P<0.01, r=-0.94).4.The expression of cIAP1mRNA in the anastomotic tissue of experimental group(25.85±6.61～36.85±4.21)were more significant compared with the control group(2.68±1.05～3.10±1.00)at all the time points (P<0.05),located in the interstitial cells, especially in fibroblasts; there was no significant difference among all the stages (P>0.05).The expression of Caspase3 in the anastomotic tissue of experimental group (2.65±0.60～4.37±0.62)were significantly less than the control group (10.15±2.06～11.96±3.69)(P<0.05),the expression located in epithelial was relatively stronger and there was no significant difference among all the phases (P>0.05).There was negative correlation between cIAP-1 mRNA and caspase3 protein in expression (P<0.01, r=-0.95).Conclusion:The activation of NF-κB and the expression of its downstream apoptosis protein cIAP-1,c-FLIP of bile duct (muscle) fibroblasts may play an important role in bile duct scar healing after injury by respectively inhibiting the activation of terminal proteins of executive apoptosis caspase3,8.
|
PDF Full Text Request