| 摘要: | 正常腸道微生物叢 (normal microbiota)的存在，可以幫助宿主抵抗病原菌的侵襲及健全免疫力，甚至提供維生素等營養素。如果腸道菌叢發生變化或被破壞，就會給與伺機性病原菌(opportunistic pathogen) 造成宿主發生腸炎或感染的機會。本實驗為釐清腸道共生菌參與宿主的免疫調節作用，採用存在大部分哺乳動物腸道中之共生菌Bacteroides fragilis (B. fragilis)，屬於絕對厭氧之格蘭氏陰性菌。以管餵方式定殖(colonization)到8至9週齡無菌小鼠(Germ-free mice, GF)，於接菌四星期後犧牲並進行分析該菌對於無菌小鼠免疫系統之影響，對照組為不含B. fragilis (BF)之無菌小鼠。結果顯示：1.比較BF實驗組與GF對照組之體重並無顯著性差異，盲腸重量及結直腸長度則具有顯著性差異；2.由血球計數器計算週邊單核細胞 ( peripheral blood mononuclear cells, PBMC)結果得知，白血球數量在BF組與GF組小鼠並無明顯差異，中性球與單核球的比例則有顯著性增加；3.定殖四星期後，於盲腸與結腸之B. fragilis菌約為1010 cfu/g；4.以流式細胞儀分析脾臟、腸系膜淋巴結與培耶氏斑的T細胞與B細胞族群，結果發現投與B. fragilis四星期後可誘發腸系膜淋巴結T細胞與B細胞的比例有顯著性增加；5.分析結直腸細胞產生細胞素的能力，結果顯示，BF組小鼠抗炎症細胞素IL-10表現量顯著高於GF小鼠； 6. in vitro培養BF組小鼠脾臟細胞經LPS或Con A刺激釋放Th1 相關TNF-α與IFN-γ等細胞素的能力比GF小鼠高，相反的其釋放Th2相關細胞素如IL-4、IL-10則比GF小鼠低。综合上述結果顯示，B. fragilis具有調節無菌小鼠免疫的功能，並可藉由管餵方式建立B. fragilis單一已知菌動物(mono-associated gnotobiotic mice) 模式。
Normal flora are bacteria, fungi, and protozoa that live on or within the bodies of animals and plants. Usually, they do not cause disease in healthy individuals. Instead, they are commensalists or mutualists with regard to the host. That is, in addition to basically not harming the host, they can even do some good for prevention opportunistic pathogens. In order to investigate the immunological roles of symbiotic bacteria on intestine mucosal, germ-free 8- to 9-week old male C57BL/6JNarl mice , were housed under sterilized isolator and with free access to food and water. Mice were oral with 0.5 ml, 1108/per of Bacteroides fragilis (B. fragilis, BF) dissolved in saline, as well as oral with 0.5 ml saline as a control group. The animals were sacrificed at 1 month for physiology, gross and histopathology, immunohistochemistry, flow cytometry, cytokine, and PCR analysis. The results show no significant difference of body weight as well as WBC between both groups. However, the neutrophils and monocytes were induced in BF group. The lymphocytes were aggregated in lamina propria of BF’s intestine. In addition, the GALT hyperplasia was also noted. Under immunohistochemistry and flow cytometry, B cells in GALT, CD3 T cells in MLN were induced by B. fragilis. The production of IL-10 was also increased in colon. The splenocytes were stimulated by LPS or ConA in vitro. Interestingly, the production of TNF-α and IFN-γ (Th1) and reduction of IL-4 and IL-10 (Th2) were induced by B. fragilis. The results suggested that B. fragilis not only stimulated differention of local immunity but also modulated the systemic immune system. The mono-associated gnotobiotic mice model was first established in Taiwan. |
