The Clinical Analysis And Pathogenesis Explore Of Children With Inflammatory Bowel Disease

Background Inflammatory bowel disease (inflammatory bowel disease, IBD), including Crohn’s disease (Crohn’s disease, CD) and ulcerative colitis (ulcerative colitis, UC), is an intestinal chronic nonspecific inflammatory disease. In recent years, with the rapid development of digestive endoscopic techniques widely used in children with chronic gastrointestinal inflammatory diseases, children of IBD diagnosis rate show significantly upward trend. But the etiology of this disease is unclear, the pathogenesis is unknown, treatment is more difficult.The present study showed that the etiology of IBD may be closely related to the infection, immune disorders and other factors. Increased intestinal permeability caused by abnormal immune reaction plays an important role in the pathogenesis of IBD. Tumor necrosis factor-a (Tumor necrosis factor-alpha, TNF-a) which mainly produced by activated macrophages are important molecules in the regulation of inflammatory immune response. Studies have found that TNF-a lead to destruction of the intestinal epithelial barrier function and increased permeability, as which is the main pathological basis of IBD. Therefore, for children with refractory IBD, early treatment with monoclonal antibodies (infliximab) of TNF-a obtained better clinical results. It was indicated that TNF-a may be one of the key factors in the development of IBD, but the specific mechanism unknown. So, depth studying the role of TNF-a in IBD help to elucidate the pathogenesis of IBD, and to provide new therapeutic targets.Object:1. To analysis the clinical features of children with inflammatory bowel disease, and to evaluate the therapeutic effect of TNF-alpha monoclonal antibody (infliximab) to IBD.2. To discuss the influence of TNF-a on the intestinal epithelial barrier permeability and the regulator mechanism.Method:1. All of the clinical data with dignosis of IBD were collected from January2007to January2012in our hospital. According to the diagnosis and treatment guidelines of IBD announced by the World Gastroenterology Organization (WGO,2010) and the diagnosis standards of CD and UC for IBD in "Chinese Consensus on Diagnosis and Treatment Standard of Inflammatory Bowel Disease", which announced by the Children IBD collaborative group, Gastroenterology group, Chinese Medical Society (2010),12cases which met the inclusion criteria were chosen to do retrospectively analyzed. 2. Caco-2cells were cultured in vitro to establish a model of intestinal epithelial barrier and divided randomly into two groups:control group and NF-κB inhibition group. The control group was transfected with no-load plasmid (vector), and NF-κB inhibition group was transfected with DN-Mu ikBa plasmid. The changes in transepithelial electrical resistance (transepithelial electrical resistance, TEER) of Caco-2cells were measured using an electrical resistance system. The changes in cytoskeleton were observed by direct staining with rhodamine-phalloidin. The activation of nuclear factor-κB (NF-κB) was determined by luciferase reporter gene assay. The phosphorylation expression of myosin light chain (MLC) was detected by immunoblotting (Western blotting).Result:1. There were3cases of Crohn’s disease (CD) and9cases of ulcerative colitis (UC) in nine cases of IBD, with8males and4females, the age of onset was6months to12years (median age of6years). Twelve cases with IBD had no positive family history, clinical manifestations were weight loss, anemia in ten cases (83.3%), abdominal pain in9cases (75.0%), fever, blood in the stool in7cases (58.3%), diarrhea in eight cases (66.7%). Laboratory tests show that CRP elevated in all of the patients, ESR elevated in10cases (83.3%), white blood cell count elevated in10cases (83.3%). Pathological results found superficial small ulcers in8cases (66.7%), chronic mucosal inflammation in seven cases (58.3%), there was no non-caseous granuloma. Nine cases were eased after giving5-amino salicylic acid or sulfasalazine, corticosteroids. Three cases recurrence, including two cases of children with CD (male), one case of child with UC (women). After application infliximab monoclonal antibody to the three recurrent cases, CD activity index was significantly reduced, the clinical symptoms in children obtained long-term remission.2. TNF-a increases the paracellular permeability of Caco-2cell. According to the TEER results, TNF-a for6hours and48hours transepithelial resistance were492±13.74(Ω· cm2) and373.67±19.55(Ω·cm2), of DNMu-IkBa group at the same time were532±17.77(Ω· cm2),455±15.39(Ω· cm2), the difference was significant (P<0.05).3. After0.5hours of TNF-alpha preconditioning the NF-KB activity was significantly increased. The activation effects of TNF-alpha to NF-KB in Caco-2cells transfected with DNMu-IkBa plasmid was significantly inhibited.4. F-actin was no significant change after TNF-alpha preconditioning in6hours. After24hours in TNF-alpha preconditioning, the peripheral contours of F-actin blurred, tends to disintegration and dissipate. Transfected DN-mulkBa plasmid in Caco-2cells was significantly reduced these changes.5. MLC-P phosphate level was significantly regulated after TNF-alpha processing6hours Caco-2cells, which reached the peak in 24hours. Therefore, after transfection the DN-mulkBa plasmid to inhibit NF-kB activity, the level of MLC-P phosphorylation was inhibited obviously.Conclusions:1. The treatment of IBD children is more difficult and easy to relapse. The use of anti-TNF-a drug infliximab treatment to refractory inflammatory bowel disease of children, especially children with CD is valid.2. TNF-α increases the paracellular permeability of Caco-2cell monolayers perhaps by inducing NF-κB activation, F-actin rearrangement and MLC phosphorylation.