The Age Distribution Characteristics Of Pediatric Intestinal Segmented Filamentous Bacterium (SFB) And Primary Exploration Of The Relationship Between SFB And Intestinal Mucosal Immunity

Background:Segmented filamentous bacterium (SFB) is a gram positive, spore producing and anaerobic bacteria, widely exists in the intestinal tract of a variety of vertebrate and invertebrate animals, presenting as a kind of specific morphological structure "original" bacteria. SFB is closely related to immune maturing of the host, promoting secretion of sIgA, regulating differentiation and balance of T cells, inducing differentiation of Th17 cell and resisting pathogenic bacterium infection. Since it is very difficult to culture for SFB in vitro, studies should be carried out through animal model with sole bacterium colonization including rat, mice, chicken, et al. It was found that SFB significantly promote secretion of intestinal sIgA and differentiation of Thl7 cell, both of which are the vital functional molecule in local intestinal mucosal immune barrier. The functions of slgA include inhibiting intestinal bacterium adhering to mucosal surface, neutralizing toxin, enzyme and virus in intestine, and sealing antigen. SFB also shows the function as degranulation to eosinophilic and basophilic granulocyte cells, and antibody dependent cell-mediated cytotoxicity (ADCC) effect. Th17 cell can protect intestinal tract surface avoiding infection from bacterium and fungus at the appropriate concentration. Besides, colonization of SFB can also regulate differentiation of Treg cell, preventing autoimmunity response induced by Th17 excessive differentiation. Multiple studies have demonstrated that the colonization of SFB in animal (like mice, rat and chicken) intestine showed an age dependent manner, however, there were few studies concerning on the colonization, distribution and its influence on the host immune system of SFB in human intestinal tract.Objective:To investigate the age distribution characteristics of pediatric intestinal SFB; and to analyse the relationship between human intestinal SFB colonization and intestinal mucosal immunity by measuring intestinal sIgA, immune-associated cells of terminal ileum mucous and Th cell differentiation associated transcription factors using immunohistochemistry methods. Methods:1. The age distribution characteristics of pediatric intestinal SFB:the non-gastrointestinal, non-infectious surgery patients were enrolled from November 2013 to January 2014, excluding applying for antibiotics and microecologics. Totally 177 patients were enrolled and their fresh faeces were collected. DN A of the faeces was extracted and SFB sequence was acquired from GenBank. The specific primer was designed according to 16SrRNA/DNA specific gene sequence of the bacterium. Furthermore, real-time PCR was used to analysis quantitatively to confirm exists of SFB, and then statistical analysis was carried out.2. Primary exploration of the relationship between SFB and intestinal mucosal immunity:the patients receiving colonoscopy but no any abnormal findings were enrolled from October 2012 to June 2014. The fresh intestinal fluid of ileocecal junction was collected and the concentration of sIgA was measured by ELISA for statistical analysis; the terminal ileum mucosa samples collected during colonoscopy were analyzed using immunohistochemistry method. The number of IL-17A cell and intraepithelial lymphocyte in terminal ileum were measured, as well as the transcription factor associated differentiation of Th1,Treg and Th17 cells, for primary exploration of the relationship between SFB and intestinal mucosal immunity.Results:1. The age distribution characteristics of pediatric intestinal SFB:SFB were positive in 34 cases among 177 patients, with positive rate of 19.2%; the positive rate of SFB varies in different age, with 40.0%,47.4%,31.6%,15.0%,11.8%,13.3%,15.4% and 3.7% in age 0-1 years,1-2 years,2-3 years,3-4 years,4-5 years,5-6 years,6-7 years and 7-15 years, respectively. Spearman relative test showed that approximate T value was -5.095, P<0.001, indicating SFB was mainly planted in the gut such as infants and young children less than 3 years old.2. Relationship between human intestinal SFB and sIgA:There were 24 and 23 cases in SFB positive and negative groups respectively, and sIgA concentration of intestinal fluid in SFB positive group increased compared with SFB negative group with significant statistical difference (P<0.05).3. Relationship between human intestinal SFB and intestinal mucosal immune cells and transcriptional factor:There were 12 and 11 cases in SFB positive and negative groups, respectively, and there was no statistical difference of lymphocyte number in terminal ileum mucosal epithelial cell between these two groups. The number of IL-17A cells in terminal ileum mucosa of SFB positive group decreased compared with SFB negative group with significant statistical difference (P<0.05). There was no statistical difference of Th1 cell differentiation-associated transcriptional factor T-bet expression, Treg cell differentiation-associated transcriptional factor FOXP3 expression and Th17 cell differentiation-associated transcriptional factor ROR-γt expression between these two groups.Conclusion:Human SFB mainly plants in intestine tract of infants and young children less than 3 years old; Human SFB promotes intestine secreting sIgA; The effect of intestinal colonization of human SFB on the children’s mucosal immune functions may be different from that of animal SFB.