Role of the bacterial gut microbiota and innate immunity in Campylobacter jejuni-enteritis: Investigations in gnotobiotic mouse models of inflammation

空肠弯曲菌引起的肠炎是世界范围内流行最广泛的腹泻疾病之一。在工业化国家，空肠弯曲菌感染的临床特征最常见的是下肠道的急性严重炎症反应，并伴有水样腹泻，包括便血、发热和腹痛。炎症是由肠黏膜内粒细胞和巨噬细胞的浸润引起的。这些先天免疫细胞触发炎症反应并持续组织破坏，导致健康粘膜呈现的肠道屏障功能严重紊乱，以抵御自然共生肠道菌群中的非致病细菌。因此，不仅空肠弯曲菌而且其他易位肠道细菌也侵入粘膜下组织部位，并与免疫细胞直接接触，从而加重炎症。提示先天免疫系统和正常肠道菌群可能通过参与炎症过程的调控，在空肠弯曲菌诱导的免疫病理过程中发挥关键作用。一些临床研究表明，在炎症性肠病患者中，正常肠道菌群的非致病性共生细菌可能是肠道炎症持续存在的关键成分。然而，到目前为止，有关空肠弯曲菌肠炎的肠道生态和先天免疫方面的研究尚未见报道。细菌分子通过与Toll样受体(TLRs)或NOD2蛋白结合来加重或抑制炎症反应，这两种受体分别在天然免疫系统的细胞如粒细胞、巨噬细胞或Paneth细胞中高水平表达。由此产生的信号转导事件启动促炎或抗炎细胞因子反应，调节包括T细胞和B细胞在内的先天免疫和获得性免疫的反应性。我们团队和其他人的实际研究表明，TLRs和NOD2在调节胃肠道的急性和慢性炎症过程中发挥着重要作用。基于这些发现，我们推测，共生细菌肠道微生物区系和先天免疫系统都有助于空肠弯曲菌肠炎的持续或抑制。我们将利用灵知生菌小鼠模型，研究肠道菌群与空肠弯曲菌定植及致病机制的相互作用，并探讨先天免疫在空肠弯曲菌肠炎中的作用。宿主先天免疫对空肠弯曲菌肠炎的影响将在TLR2、TLR4、TLR9或NOD2基因以及信号转导蛋白MyD88或TRIF编码基因明确缺陷的小鼠中进行研究。相应的机制将在体外进一步表征。空肠定植和肠道炎症将在携带特定肠道细菌或受到细菌化合物挑战的诺生菌小鼠身上进行研究。随后将使用纯化的TLR和/或NOD2配体对相应的动物进行治疗，以满足Kochs的假设，并评估TLR/NOD激动剂/拮抗剂、特定的肠道细菌或肠道菌群调节是否适合在空肠炎过程中抑制免疫激活和减少急性炎症反应。最后，我们的目标是在我们已建立的回肠炎和结肠炎小鼠模型中，确定肠道炎症反应对对空肠弯曲菌表现出的定植抵抗的影响。

Enteritis caused by C. jejuni is one of the most widespread diarrhoeal diseases worldwide. In industrialized nations, the clinical spectrum of C. jejuni-infection is most commonly characterized by acute severe inflammatory responses in the lower intestinal tract accom-panied by watery diarrhoea including bloody stools, fever and abdominal pain. The inflam-mation is initiated by infiltration of the intestinal mucosa with granulocytes and macropha-ges. These innate immune cells trigger inflammatory responses and perpetuate tissue de-struction leading to a severe disturbance of the intestinal barrier functions presented by the healthy mucosa to defend against non-pathogenic bacteria of the natural commensal inte-stinal flora. In consequence, not only C. jejuni but also other translocating gut bacteria infil-trate submucosal tissues sites and are coming in direct contact with immune cells thereby aggravating inflammation. This suggests that the innate immune system and the normal gut flora might play a key role in C. jejuni-induced immunopathology by taking part in the modulation of the inflammatory process. Several clinical studies have shown that in patients suffering from inflammatory bowel diseases non-pathogenic commensal bacteria of the normal intestinal flora might represent critical components in the perpetuation of intestinal inflammation. However, such aspects of intestinal ecology and innate immunity have not been studied in C. jejuni-enteritis so far. Bacterial molecules aggravate or sup-press inflammatory responses by binding to Toll-like receptors (TLRs) or the NOD2-pro-tein, both of which are expressed at high levels by cells of the innate immune system such as granulocytes, macrophages or Paneth cells, respectively. The resulting signal transduc-tion events initiate pro-inflammatory or anti-inflammatory cytokine responses modulating the reactivity of innate and adaptive immunity, including T- and B-cells. Actual research in our group and others has demonstrated that TLRs and NOD2 play important roles in the modulation of acute and chronic inflammatory processes in the gastrointestinal tract. Based upon these findings we hypothesize that both, the commensal bacterial gut micro-flora and the innate immune system, contribute to the perpetuation or suppression of C. jejuni-enteritis. We will use gnotobiotic mouse models to investigate the interplay of the intestinal microflora with C. jejuni colonization and pathogenesis and to study the role of innate immunity in C. jejuni-enteritis. The influences of host innate immunity on C. jejuni-enteritis will be studied in mice with well-defined defects in the TLR2, TLR4, TLR9, or NOD2 genes as well as genes coding for the signal-transducing proteins MyD88 or TRIF. The corres-ponding mechanisms will be further characterized in vitro. C. jejuni-colonization and intestinal inflammation will be investigated in gnotobiotic mice carrying defined gut bacteria or challenged by bacterial compounds. A subsequent treatment of the corresponding animals with purified TLR- and/or NOD2-ligands will be used in order to fulfil the Koch`s postulates and to evaluate if TLR-/NOD-agonists/ antagonists, defined gut bacteria, or modulation of the intestinal flora might be suitable to suppress immune activation and reduce acute inflammatory responses in the course of C. jejuni-enteritis. Finally, we aim to determine the influences of intestinal inflammatory responses on colonization resistance displayed against C. jejuni in our well-established mouse models for ileitis and colitis.