耐药结核分枝杆菌感染的免疫治疗及艰难梭状芽孢杆菌毒素的结构域功能与病理研究
|
摘要
结核分枝杆菌(Mycobacterium tuberculosis)和艰难梭状芽孢杆菌(Clostridium difficile)都是重要的威胁公众健康的感染型病原微生物。本文主要针对耐药结核分枝杆菌感染致病的免疫治疗和艰难梭状芽孢杆菌毒素蛋白的结构域及病理机制展开了研究。
结核分枝杆菌在全球广泛流行；在我国,结核病是第一大流行性传染病之一。当前,结核病治疗面临的最严峻挑战在于耐药结核菌株特别是多重耐药菌株的出现。由于现有药物治疗方案无法完全清除各种耐药菌株,致使对传染源的控制难度增大,因此,亟需发展新的结核病治疗方案。采用细胞因子调节宿主对入侵病原菌产生免疫应答的免疫治疗法是可能成为临床治疗结核病的潜在有效策略之一。本研究在结核病小鼠模型中评估了细胞因子IL-2、GM-CSF与异烟肼(INH)、利福平(RIF)联合使用治疗结核菌感染的治疗效果。我们首先在药物敏感型结核杆菌H37Rv感染小鼠模型中,通过测定小鼠肺部和脾脏菌落数和分析肺部组织病理程度筛选有效的治疗方案。和未治疗组小鼠对比,经过IL-2或GM-CSF治疗后的小鼠肺部的菌落数分别减少了0.82Log10(P<0.01)和0.58Log10(<0.05),而脾脏的菌落数分别减少了1.42Log10(P<0.01)和1.22Log10(P<0.01)。虽然当单一细胞因子与INH+RIF (HR)联合使用时,并未在抑菌率上表现出任何优势,但是肺部组织病理切片显示联合免疫治疗组小鼠肺部损伤明显减弱。随后,本研究在耐药结核杆菌OB35感染小鼠模型中,通过监测小鼠体重变化、肺部和脾脏菌落计数、以及肺部病理变化评测了免疫治疗法对耐药结核病的治疗效果。和未治疗组比较,免疫治疗能有效降低肺脾菌落数。虽然化学药物HR治疗后小鼠体内菌落数也有所下降,但是只有经免疫治疗后的小鼠肺部才表现较弱的病理性损伤。值得一提的是两种细胞因子共同参与的免疫治疗能比化学药物HR治疗更进一步降低体内菌落数,其中肺部CFU进一步减少1.02Log10(P<0.01),而脾脏CFU进一步减少1.34Log10(P<0.01)。同时,通过对平均体重、存活率以及组织病理程度的分析,我们发现IL-2和GM-CSF共同参与免疫治疗具有协同效应,可有效增强一线药物治疗耐多药结核杆菌感染的效果。
艰难梭状芽孢杆菌在发达国家是抗生素治疗引起的院内感染型腹泻的主要原因之一。由于日益增加的发病率和死亡率,艰难梭菌感染(CDI)在全世界已引起了广泛关注。近来出现的高毒耐药菌株更是急剧增加了CDI的发病率,使CDI的治疗和控制面临更严峻的挑战。现阶段对CDI的病理仍然不是很清楚。目前的研究一直认为,从属于大梭菌毒素家族的两个外毒素,毒素A (TcdA)和毒素B (TcdB)是引起疾病的主要原因,但是它们的致病机制以及各自在疾病中的作用意义并不十分清楚。TcdA和TcdB都是多结构域蛋白,主要通过糖基化胞内的Rho GTPase引起宿主细胞中毒。迄今为止发现的毒素功能结构区至少有四个,包括：葡萄糖基转移酶区(GT)、半胱氨酸水解酶区(CPD)、跨膜区(TD)以及受体结合区(RBD)。这四个结构区的功能和致病机制目前尚未完全明确。特别是跨膜区作为毒素最大的结构功能区,涵盖了毒素几乎50%的氨基酸序列,但是至今还未充分阐明其确切的生物学功能和分子机制。基于上述背景,本论文关于艰难梭状杆菌毒素的研究重点涵盖了两大部分：1、艰难梭菌TcdA和TcdB的结构区功能研究。2、TcdA和TcdB致病机制。在第一部分中,我们重点研究了艰难梭菌两个毒素的跨膜区和受体结合区在毒素信号转导通路和疾病发生发展中的生物学功能。我们发现位于跨膜区C端的一段97个氨基酸的片段(D97)对TcdB的细胞毒性起着十分重要的作用。将这个片段的编码序列从TcdB的基因中去除后,表达并纯化得到
个截短的毒素蛋白,命名为TxB-D97。实验发现D97片段的缺失并没有影响TcdB蛋白的其它结构功能区的功能,例如,突变毒素和野生型毒素与细胞的结合以及被细胞胞吞的情况基本相似；而且它们在六磷酸肌醇(Insp6)作用下释放GT区效率也一样,同时GT区的糖基转移酶活性在体外的无细胞体系中也表现一致。尽管如此,TxB-D97的细胞毒性和细胞致病性与野生型TcdB相比减弱了至少105。与野生型TcdB不同,TxB-D97不能有效将活细胞中Rac1蛋白糖基化,也不能刺激巨噬细胞分泌TNF-α,而刺激巨噬细胞分泌TNF-α的特性是与细胞质中毒素GT的活性密切相关。同时,用野生型TcdB和突变型毒素TxB-D97分别处理细胞后,用免疫共沉淀方法无法在TxB-D97处理细胞的细胞抽提物中检测到有GT释放。上述实验结果证明位于毒素跨膜区C端紧靠RBD的片段D97是TcdB将GT区运送到细胞质中不可缺少的区域。除D97外,在毒素结构功能区的研究中,为了定义毒素肠毒性与毒素结构区的关系,我们通过交换两个毒素的GT、RBD结构区构建了嵌合体毒素。在一个已成熟使用的小鼠回肠封闭实验模型中,我们严格检查了嵌合体毒素的肠毒性,并对肠道的积水进行了定量分析,检测了炎症细胞因子的诱导分泌,中性粒细胞渗出,肠上皮组织损伤。通过对这些数据的分析我们定义了嵌合体毒素的肠毒性。嵌合体TxdA-Br安装有TcdB的RBD结构区,它在小鼠回肠节中引起了肠组织损伤、大量中性粒细胞渗出,炎症细胞因子表达以及肠积水。而另一个嵌合体TxdB-Ar虽然安装了TcdA的RBD结构区却不能诱发所有肠毒性反应。此外,糖基转移酶活性缺失的突变型毒素aTcdA也基本丢失了其肠毒的活性。这些数据表明TcdA的肠毒性不是由它的RBD结构区决定。在艰难梭状芽孢杆菌毒素第二部分的研究中,我们探讨了TcdA和TcdB在CDI发病机制中的不同作用。在这部分实验中,我们建立了一个巨大芽孢杆菌(Bacillus megaterium)小鼠口服模型。利用巨大芽孢杆菌表达单一毒素来替代艰难梭菌,通过灌胃法,使毒素在这种非致病细菌的保护下到达小鼠胃肠道。利用这个实验模型,我们发现表达TcdB的巨大芽孢杆菌,不依赖于TcdA或其它来源于艰难梭菌的辅助因子,能引起小鼠发病和死亡。而TcdB不仅在肠道引起病变而且还引起了肺部严重损伤,这可能与TcdB进入了血液循环表现出全身体循环毒性有关。相比TcdB, TcdA以乎是一个更专一型的肠毒素,它虽然也能进入全身体循环,但是TcdA只在肠道诱发严重病变和炎症反应。从表达TcdA的巨大芽孢杆菌灌胃的小鼠体内收集的血清和胸积水中,我们检测到促炎症细胞因子IL-1β、IL-6、KC和TNF-α的表达水平都显著升高,这充分说明TcdA是一个高炎症性的毒素。
综上所述,本研究通过多种分子生物学、免疫学和病理学方法,评价耐药与非耐药结核杆菌感染的细胞因子免疫治疗方案,研究艰难梭菌TcdA和TcdB的结构区的功能,并探讨TcdA及TcdB在疾病病理中的作用。针对结核分支杆菌感染的免疫治疗我们得出初步结论：IL-2和GM-CSF是潜在能提高一线抗结核药物效力的免疫调节剂；同时多个细胞因子参与的免疫治疗将可能成为有效控制耐药结核病的潜在治疗策略。而在艰难梭状杆菌毒素的研究中,我们发现(1)位于TcdB跨膜区C端的一段97个氨基酸的片段(D97)对艰难梭菌TcdB的GT运输到目的细胞细胞质的过程有非常重要的作用；(2)TcdA和TcdB的受体结合区(RBD)不是决定肠毒性的主要功能区,而GT的活性是肠毒性的必要条件之一；(3)TcdA和TcdB分别能独立在体外和体内引起上皮细胞屏障功能障碍,从而增加了细胞旁通路物质流和毒素转运；(4)虽然TcdB不是严格定义上的肠毒素,但是不依赖于TcdA或其它艰难梭菌产生的辅助因子,TcdB也能在体内引起疾病；(5)TcdA和TcdB都能进入血液循环,导致血毒症和严重甚至致命的系统性疾病；(6)与TcdA相比,TcdB炎症性更弱；(7)TcdB是一个毒性强大的体循环系统毒素,能引起肠道和全身组织损伤。这些研究结果填补了艰难梭状芽孢杆菌研究领域的一些知识空白,为进一步了解该微生物毒素蛋白在体内的病理模型奠定了基础。
Mycobacterium tuberculosis and Clostridium difficile are both infectious bacteria threatening public health. In this study, we are focusing on the immunotherapy of M. tubercosis infection and the domain functions and pathogenesis of C. difficile toxins.
Tuberculosis (TB) is prevalent globally. In China, it is one of the most threatening epidemic diseases. The emergence of multi-drug resistant TB (MDR-TB) makes it difficult to treat TB because the current regimens failed to clear up MDR-TB in patients. New therapeutic regimens to treat TB are urgently needed due to the emergence of multidrug resistant tuberculosis (MDR-TB). Cytokine-based regulation of host immune response is likely to be one of the available methods for clinical treatment of tuberculosis. In this study, we investigated the therapeutic effects of IL-2and GM-CSF co-administrated with INH (isoniazid) and RIP (rifampin) to treat TB in mouse model of TB infection. We first evaluated the effective regimens in drug susceptible M. tuberculosis H37Rv infected mice based on bacterial counts in lungs and spleens and the degree of histopathology in the lungs. Compared with untreated group, both IL-2and GM-CSF monotherapy reduced the bacterial number in the lungs by0.82(P<0.01) and0.58(P<0.05) log10respectively while in spleens by1.42(P<0.01) and1.22(P<0.01) log10respectively. Although the combination of chemical drugs and single cytokine regimens in mice did not show any advantage in terms of reducing the CFU, the lungs had much less lesions compared with the chemotherapy alone group. Next, the effects of immunotherapy regimens were assessed for MDR-TB strain OB35infected mice by detecting mean body weight and survival, bacterial counts in lungs and spleens, and histopathological changes in the lungs. Compared with untreated control and chemotherapy alone groups, immunotherapy with either cytokine had a significant reduction of CFU in lungs and spleens. The pathological changes of lungs from immunotherapy groups were also less severe and the lesions were limited compared with that of the control mice. Notably, the chemotherapy in combination with both cytokines group (IR-IL-2-GM-CSF) reduced the lung and spleen bacterial loads by1.02,1.34log10, respectively compared with chemotherapy alone. Additionally, the average body weights, survival rates, and histopathological results also revealed the synergistic effect of IL-2and GM-CSF when co-administrated with INH-RIF (HR).
Clostridium difficile is one of the leading causes of nosocomial diarrhea associated with antibiotic treatment in developed countries. The increasing morbidity and mortality of C. difficile infection (CDI) have attracted more attentions globally. The emergence of hypervirulent antibiotic-resistant strains contributes the recently sharp increase of incidence and severity of CDI. The pathogenesis of CDI is not well-understood. Two exotoxins, TcdA and TcdB, which belong to large clostridia toxin family, are thought to be the main causes of disease. Both toxins are multi-domain proteins and intoxicate target cells by glucosylating Rho GTPases. The toxins consist of at least4functional domains including a glucosyltransferase (GT) domain, a cystine protease domain (CPD), a transmembrane domain (TD), and a receptor binding domain (RBD). The functions of these molecular domains and their roles in pathogenesis of disease are not fully understood; especially the TD, which is the largest segment of toxins and comprises nearly50%of the protein, is rarely investigated. There are two research objectives in this study:to investigate domain function and to determine toxins'role in the pathogenesis of the disease. As for the first objective, we investigated the functions of toxin domains in their molucular action and disease manifestation. We identified that a97-amino-acid fragment (D97), located in the C-terminus of the TD, is essential for the cellular activity of TcdB. We deleted D97and expressed a truncated protein (designated TxB-D97), and found that the deletion did not appear to affect the functions of the other domains. The cell binding and uptake were similar between the mutant and wild type TcdB. Both wild type and mutant toxins released their GT domains similarly in the presence of inositol hexakisphosphate (Insp6), and showed a similar intrinsic glucosyltransferase activity in a cell-free glucosylating assay. Despite these, the cytopathic and cytotoxic activities of TxB-D97were reduced by more than5logs compared to wild type toxin and the mutant toxin failed to glucosylate Rho GTPase Racl of intact cells. Unlike wild type TcdB, the mutant toxin failed to induce macrophages to produce tumor necrosis factor alpha (TNF-a), an outcome dependent on the GT activity of the toxin. Cellular fractionation demonstrated that the TxB-D97was unable to liberate its GT domain efficiently into cytosol. Our data demonstrated that the D97fragment, located in the C-terminus of the TD, adjacent to the RBD, is essential for the delivery of the GT domain into the cell cytosol. In order to define the enterotoxicity of the toxins within their molecular domain(s), we generated several chimeric toxins by switching domains between the two toxins. We examined the enterotoxicity of those chimeric toxins in a well-established mouse ileal loop model. The enterotoxicity of these chimeric toxins was defined by their abilities to cause fluid accumulation, induce the production of proinflammatory cytokines and influx of neutrophils, and destruct intestinal epithelia. Chimera TxdA-Br with the RBD of TcdB induced mouse intestinal tissue damage, numerous neutrophil infiltration, inflammatory cytokine production and fluid accumulation in ileum loops whereas chimera TxdB-Ar with the RBD of TcdA failed to induced any enterotoxic response. Meanwhile, glucosyltransferase-deficient mutant TcdA was essentially lost their enterotoxicity. These data indicate that the enterotoxicity of TcdA is not determined by its RBD. For the second research objective, we explored the differential role of TcdA and TcdB in CDI pathogenesis. We established a Bacillus megaterium oral challenged mouse model. B. megaterium was used as a surrogate host to express individual toxins, allowing their delivery into mouse gastrointestinal tracts. Utilizing this model, we found that B. megaterium expressing TcdB cause disease and death independent of TcdA or any other cofactors from C. difficile. TcdB caused severe lesions not only in intestinal tissues but also in lung, which is likely directly caused by systemic TcdB after its liberation into circulation. Compared to TcdB, TcdA appeared to be a more potent enterotoxin by inducing severe lesions and inflammatory response in intestine, although it can also liberate into circulation. Pro-inflammatory cytokines IL-1, IL-6, KC and TNF-a were significantly elevated in the serum and pleural fluid of mouse challenged with B. megaterium expressing TcdA, suggesting this toxin is highly pro-inflammatory.
Conclusions:For the study of immunotherapy of Mycobacterium tuberculosis infection, we conclude that IL-2and GM-CSF are potential immunomodulators to enhance the effects of chemical drugs to treat MDR-TB, which suggests that the immunotherapy with combination cytokines is likely to be a promising strategy for tuberculosis treatment in the future. For the study of Clostridium difficile toxin study, we found that:(1) D97is essential for delievery of GT domain of TcdB into cytosol of target cells;(2) RBD was not responsible for enterotoxicity and GT was required for enterotoxicity;(3) TcdA and TcdB alone caused epithelial barrier dysfunction both in vivo and in vitro, subsequently increased paracellular flux and toxin translocation;(4) Although TcdB was not an exactly defined enterotoxin, independent of TcdA or other factors from C. difficile, TcdB caused disease in vivo;(5) Both toxins liberated to circulation, resulting in toxemia and severe and fatal systemic disease;(6) TcdB is less pro-inflammatory in comparison to TcdA;(7) TcdB have potent systemic toxicity and is able to cause both intestinal and systemic destructions. These results are useful to further understand the pathogenesis of these toxins in vivo and supply new evidences for study of pathological model.
结核分枝杆菌在全球广泛流行；在我国,结核病是第一大流行性传染病之一。当前,结核病治疗面临的最严峻挑战在于耐药结核菌株特别是多重耐药菌株的出现。由于现有药物治疗方案无法完全清除各种耐药菌株,致使对传染源的控制难度增大,因此,亟需发展新的结核病治疗方案。采用细胞因子调节宿主对入侵病原菌产生免疫应答的免疫治疗法是可能成为临床治疗结核病的潜在有效策略之一。本研究在结核病小鼠模型中评估了细胞因子IL-2、GM-CSF与异烟肼(INH)、利福平(RIF)联合使用治疗结核菌感染的治疗效果。我们首先在药物敏感型结核杆菌H37Rv感染小鼠模型中,通过测定小鼠肺部和脾脏菌落数和分析肺部组织病理程度筛选有效的治疗方案。和未治疗组小鼠对比,经过IL-2或GM-CSF治疗后的小鼠肺部的菌落数分别减少了0.82Log10(P<0.01)和0.58Log10(<0.05),而脾脏的菌落数分别减少了1.42Log10(P<0.01)和1.22Log10(P<0.01)。虽然当单一细胞因子与INH+RIF (HR)联合使用时,并未在抑菌率上表现出任何优势,但是肺部组织病理切片显示联合免疫治疗组小鼠肺部损伤明显减弱。随后,本研究在耐药结核杆菌OB35感染小鼠模型中,通过监测小鼠体重变化、肺部和脾脏菌落计数、以及肺部病理变化评测了免疫治疗法对耐药结核病的治疗效果。和未治疗组比较,免疫治疗能有效降低肺脾菌落数。虽然化学药物HR治疗后小鼠体内菌落数也有所下降,但是只有经免疫治疗后的小鼠肺部才表现较弱的病理性损伤。值得一提的是两种细胞因子共同参与的免疫治疗能比化学药物HR治疗更进一步降低体内菌落数,其中肺部CFU进一步减少1.02Log10(P<0.01),而脾脏CFU进一步减少1.34Log10(P<0.01)。同时,通过对平均体重、存活率以及组织病理程度的分析,我们发现IL-2和GM-CSF共同参与免疫治疗具有协同效应,可有效增强一线药物治疗耐多药结核杆菌感染的效果。
艰难梭状芽孢杆菌在发达国家是抗生素治疗引起的院内感染型腹泻的主要原因之一。由于日益增加的发病率和死亡率,艰难梭菌感染(CDI)在全世界已引起了广泛关注。近来出现的高毒耐药菌株更是急剧增加了CDI的发病率,使CDI的治疗和控制面临更严峻的挑战。现阶段对CDI的病理仍然不是很清楚。目前的研究一直认为,从属于大梭菌毒素家族的两个外毒素,毒素A (TcdA)和毒素B (TcdB)是引起疾病的主要原因,但是它们的致病机制以及各自在疾病中的作用意义并不十分清楚。TcdA和TcdB都是多结构域蛋白,主要通过糖基化胞内的Rho GTPase引起宿主细胞中毒。迄今为止发现的毒素功能结构区至少有四个,包括：葡萄糖基转移酶区(GT)、半胱氨酸水解酶区(CPD)、跨膜区(TD)以及受体结合区(RBD)。这四个结构区的功能和致病机制目前尚未完全明确。特别是跨膜区作为毒素最大的结构功能区,涵盖了毒素几乎50%的氨基酸序列,但是至今还未充分阐明其确切的生物学功能和分子机制。基于上述背景,本论文关于艰难梭状杆菌毒素的研究重点涵盖了两大部分：1、艰难梭菌TcdA和TcdB的结构区功能研究。2、TcdA和TcdB致病机制。在第一部分中,我们重点研究了艰难梭菌两个毒素的跨膜区和受体结合区在毒素信号转导通路和疾病发生发展中的生物学功能。我们发现位于跨膜区C端的一段97个氨基酸的片段(D97)对TcdB的细胞毒性起着十分重要的作用。将这个片段的编码序列从TcdB的基因中去除后,表达并纯化得到
个截短的毒素蛋白,命名为TxB-D97。实验发现D97片段的缺失并没有影响TcdB蛋白的其它结构功能区的功能,例如,突变毒素和野生型毒素与细胞的结合以及被细胞胞吞的情况基本相似；而且它们在六磷酸肌醇(Insp6)作用下释放GT区效率也一样,同时GT区的糖基转移酶活性在体外的无细胞体系中也表现一致。尽管如此,TxB-D97的细胞毒性和细胞致病性与野生型TcdB相比减弱了至少105。与野生型TcdB不同,TxB-D97不能有效将活细胞中Rac1蛋白糖基化,也不能刺激巨噬细胞分泌TNF-α,而刺激巨噬细胞分泌TNF-α的特性是与细胞质中毒素GT的活性密切相关。同时,用野生型TcdB和突变型毒素TxB-D97分别处理细胞后,用免疫共沉淀方法无法在TxB-D97处理细胞的细胞抽提物中检测到有GT释放。上述实验结果证明位于毒素跨膜区C端紧靠RBD的片段D97是TcdB将GT区运送到细胞质中不可缺少的区域。除D97外,在毒素结构功能区的研究中,为了定义毒素肠毒性与毒素结构区的关系,我们通过交换两个毒素的GT、RBD结构区构建了嵌合体毒素。在一个已成熟使用的小鼠回肠封闭实验模型中,我们严格检查了嵌合体毒素的肠毒性,并对肠道的积水进行了定量分析,检测了炎症细胞因子的诱导分泌,中性粒细胞渗出,肠上皮组织损伤。通过对这些数据的分析我们定义了嵌合体毒素的肠毒性。嵌合体TxdA-Br安装有TcdB的RBD结构区,它在小鼠回肠节中引起了肠组织损伤、大量中性粒细胞渗出,炎症细胞因子表达以及肠积水。而另一个嵌合体TxdB-Ar虽然安装了TcdA的RBD结构区却不能诱发所有肠毒性反应。此外,糖基转移酶活性缺失的突变型毒素aTcdA也基本丢失了其肠毒的活性。这些数据表明TcdA的肠毒性不是由它的RBD结构区决定。在艰难梭状芽孢杆菌毒素第二部分的研究中,我们探讨了TcdA和TcdB在CDI发病机制中的不同作用。在这部分实验中,我们建立了一个巨大芽孢杆菌(Bacillus megaterium)小鼠口服模型。利用巨大芽孢杆菌表达单一毒素来替代艰难梭菌,通过灌胃法,使毒素在这种非致病细菌的保护下到达小鼠胃肠道。利用这个实验模型,我们发现表达TcdB的巨大芽孢杆菌,不依赖于TcdA或其它来源于艰难梭菌的辅助因子,能引起小鼠发病和死亡。而TcdB不仅在肠道引起病变而且还引起了肺部严重损伤,这可能与TcdB进入了血液循环表现出全身体循环毒性有关。相比TcdB, TcdA以乎是一个更专一型的肠毒素,它虽然也能进入全身体循环,但是TcdA只在肠道诱发严重病变和炎症反应。从表达TcdA的巨大芽孢杆菌灌胃的小鼠体内收集的血清和胸积水中,我们检测到促炎症细胞因子IL-1β、IL-6、KC和TNF-α的表达水平都显著升高,这充分说明TcdA是一个高炎症性的毒素。
综上所述,本研究通过多种分子生物学、免疫学和病理学方法,评价耐药与非耐药结核杆菌感染的细胞因子免疫治疗方案,研究艰难梭菌TcdA和TcdB的结构区的功能,并探讨TcdA及TcdB在疾病病理中的作用。针对结核分支杆菌感染的免疫治疗我们得出初步结论：IL-2和GM-CSF是潜在能提高一线抗结核药物效力的免疫调节剂；同时多个细胞因子参与的免疫治疗将可能成为有效控制耐药结核病的潜在治疗策略。而在艰难梭状杆菌毒素的研究中,我们发现(1)位于TcdB跨膜区C端的一段97个氨基酸的片段(D97)对艰难梭菌TcdB的GT运输到目的细胞细胞质的过程有非常重要的作用；(2)TcdA和TcdB的受体结合区(RBD)不是决定肠毒性的主要功能区,而GT的活性是肠毒性的必要条件之一；(3)TcdA和TcdB分别能独立在体外和体内引起上皮细胞屏障功能障碍,从而增加了细胞旁通路物质流和毒素转运；(4)虽然TcdB不是严格定义上的肠毒素,但是不依赖于TcdA或其它艰难梭菌产生的辅助因子,TcdB也能在体内引起疾病；(5)TcdA和TcdB都能进入血液循环,导致血毒症和严重甚至致命的系统性疾病；(6)与TcdA相比,TcdB炎症性更弱；(7)TcdB是一个毒性强大的体循环系统毒素,能引起肠道和全身组织损伤。这些研究结果填补了艰难梭状芽孢杆菌研究领域的一些知识空白,为进一步了解该微生物毒素蛋白在体内的病理模型奠定了基础。
Mycobacterium tuberculosis and Clostridium difficile are both infectious bacteria threatening public health. In this study, we are focusing on the immunotherapy of M. tubercosis infection and the domain functions and pathogenesis of C. difficile toxins.
Tuberculosis (TB) is prevalent globally. In China, it is one of the most threatening epidemic diseases. The emergence of multi-drug resistant TB (MDR-TB) makes it difficult to treat TB because the current regimens failed to clear up MDR-TB in patients. New therapeutic regimens to treat TB are urgently needed due to the emergence of multidrug resistant tuberculosis (MDR-TB). Cytokine-based regulation of host immune response is likely to be one of the available methods for clinical treatment of tuberculosis. In this study, we investigated the therapeutic effects of IL-2and GM-CSF co-administrated with INH (isoniazid) and RIP (rifampin) to treat TB in mouse model of TB infection. We first evaluated the effective regimens in drug susceptible M. tuberculosis H37Rv infected mice based on bacterial counts in lungs and spleens and the degree of histopathology in the lungs. Compared with untreated group, both IL-2and GM-CSF monotherapy reduced the bacterial number in the lungs by0.82(P<0.01) and0.58(P<0.05) log10respectively while in spleens by1.42(P<0.01) and1.22(P<0.01) log10respectively. Although the combination of chemical drugs and single cytokine regimens in mice did not show any advantage in terms of reducing the CFU, the lungs had much less lesions compared with the chemotherapy alone group. Next, the effects of immunotherapy regimens were assessed for MDR-TB strain OB35infected mice by detecting mean body weight and survival, bacterial counts in lungs and spleens, and histopathological changes in the lungs. Compared with untreated control and chemotherapy alone groups, immunotherapy with either cytokine had a significant reduction of CFU in lungs and spleens. The pathological changes of lungs from immunotherapy groups were also less severe and the lesions were limited compared with that of the control mice. Notably, the chemotherapy in combination with both cytokines group (IR-IL-2-GM-CSF) reduced the lung and spleen bacterial loads by1.02,1.34log10, respectively compared with chemotherapy alone. Additionally, the average body weights, survival rates, and histopathological results also revealed the synergistic effect of IL-2and GM-CSF when co-administrated with INH-RIF (HR).
Clostridium difficile is one of the leading causes of nosocomial diarrhea associated with antibiotic treatment in developed countries. The increasing morbidity and mortality of C. difficile infection (CDI) have attracted more attentions globally. The emergence of hypervirulent antibiotic-resistant strains contributes the recently sharp increase of incidence and severity of CDI. The pathogenesis of CDI is not well-understood. Two exotoxins, TcdA and TcdB, which belong to large clostridia toxin family, are thought to be the main causes of disease. Both toxins are multi-domain proteins and intoxicate target cells by glucosylating Rho GTPases. The toxins consist of at least4functional domains including a glucosyltransferase (GT) domain, a cystine protease domain (CPD), a transmembrane domain (TD), and a receptor binding domain (RBD). The functions of these molecular domains and their roles in pathogenesis of disease are not fully understood; especially the TD, which is the largest segment of toxins and comprises nearly50%of the protein, is rarely investigated. There are two research objectives in this study:to investigate domain function and to determine toxins'role in the pathogenesis of the disease. As for the first objective, we investigated the functions of toxin domains in their molucular action and disease manifestation. We identified that a97-amino-acid fragment (D97), located in the C-terminus of the TD, is essential for the cellular activity of TcdB. We deleted D97and expressed a truncated protein (designated TxB-D97), and found that the deletion did not appear to affect the functions of the other domains. The cell binding and uptake were similar between the mutant and wild type TcdB. Both wild type and mutant toxins released their GT domains similarly in the presence of inositol hexakisphosphate (Insp6), and showed a similar intrinsic glucosyltransferase activity in a cell-free glucosylating assay. Despite these, the cytopathic and cytotoxic activities of TxB-D97were reduced by more than5logs compared to wild type toxin and the mutant toxin failed to glucosylate Rho GTPase Racl of intact cells. Unlike wild type TcdB, the mutant toxin failed to induce macrophages to produce tumor necrosis factor alpha (TNF-a), an outcome dependent on the GT activity of the toxin. Cellular fractionation demonstrated that the TxB-D97was unable to liberate its GT domain efficiently into cytosol. Our data demonstrated that the D97fragment, located in the C-terminus of the TD, adjacent to the RBD, is essential for the delivery of the GT domain into the cell cytosol. In order to define the enterotoxicity of the toxins within their molecular domain(s), we generated several chimeric toxins by switching domains between the two toxins. We examined the enterotoxicity of those chimeric toxins in a well-established mouse ileal loop model. The enterotoxicity of these chimeric toxins was defined by their abilities to cause fluid accumulation, induce the production of proinflammatory cytokines and influx of neutrophils, and destruct intestinal epithelia. Chimera TxdA-Br with the RBD of TcdB induced mouse intestinal tissue damage, numerous neutrophil infiltration, inflammatory cytokine production and fluid accumulation in ileum loops whereas chimera TxdB-Ar with the RBD of TcdA failed to induced any enterotoxic response. Meanwhile, glucosyltransferase-deficient mutant TcdA was essentially lost their enterotoxicity. These data indicate that the enterotoxicity of TcdA is not determined by its RBD. For the second research objective, we explored the differential role of TcdA and TcdB in CDI pathogenesis. We established a Bacillus megaterium oral challenged mouse model. B. megaterium was used as a surrogate host to express individual toxins, allowing their delivery into mouse gastrointestinal tracts. Utilizing this model, we found that B. megaterium expressing TcdB cause disease and death independent of TcdA or any other cofactors from C. difficile. TcdB caused severe lesions not only in intestinal tissues but also in lung, which is likely directly caused by systemic TcdB after its liberation into circulation. Compared to TcdB, TcdA appeared to be a more potent enterotoxin by inducing severe lesions and inflammatory response in intestine, although it can also liberate into circulation. Pro-inflammatory cytokines IL-1, IL-6, KC and TNF-a were significantly elevated in the serum and pleural fluid of mouse challenged with B. megaterium expressing TcdA, suggesting this toxin is highly pro-inflammatory.
Conclusions:For the study of immunotherapy of Mycobacterium tuberculosis infection, we conclude that IL-2and GM-CSF are potential immunomodulators to enhance the effects of chemical drugs to treat MDR-TB, which suggests that the immunotherapy with combination cytokines is likely to be a promising strategy for tuberculosis treatment in the future. For the study of Clostridium difficile toxin study, we found that:(1) D97is essential for delievery of GT domain of TcdB into cytosol of target cells;(2) RBD was not responsible for enterotoxicity and GT was required for enterotoxicity;(3) TcdA and TcdB alone caused epithelial barrier dysfunction both in vivo and in vitro, subsequently increased paracellular flux and toxin translocation;(4) Although TcdB was not an exactly defined enterotoxin, independent of TcdA or other factors from C. difficile, TcdB caused disease in vivo;(5) Both toxins liberated to circulation, resulting in toxemia and severe and fatal systemic disease;(6) TcdB is less pro-inflammatory in comparison to TcdA;(7) TcdB have potent systemic toxicity and is able to cause both intestinal and systemic destructions. These results are useful to further understand the pathogenesis of these toxins in vivo and supply new evidences for study of pathological model.
引文
[1]Report WHO (2009) Global Tuberculosis Control:Epidemiology, strategy, financing.
[2]彭卫生,王英年,肖成志等.新编结核病学.第二版.北京：中国医药科技出版社,2003.
[3]黄晓燕,严杰,李淑萍.结核分枝杆菌感染细胞的信号传导通路研究进展,中国生物制品学杂志,2006,19(3)：323-325.
[4]肖和平.结核病防治新进展.上海：复旦大学出版社,2004.
[5]谢惠安,肖成志,宋礼章.结核病化学疗法与化疗实施.北京：化学工业出版社,2005.
[6]Current development and future prospects in chemotherapy of tuberculosis
[7]王莉.浅谈细胞因子的网络性.河北省科学院学报,2002,19：92-97.
[8]唐神结,肖和平.细胞因子网络和结核病.中华结核和呼吸杂志,2001,24(9)：571-574.
[9]Pellegrini P, Contasta I, Berghella AM, et al. The TH1 and TH2 cytokine network in healtyh subjects:suggestions for experimental studies to create prognostic and diagnostic indices for biotherapeutic treatments[J]. Cancer Biotherapy & Radiopharmaceuticals.2000, 15:267-278.
[10]Contasta I, Pellegrini P, Berghella AM, et al. Cell cycle control in cellular homeostasis during the immune response interactions betweenTH1, TH2 cytokines, and Bcl2 and p53 molecules[J]. Cancer Biotherapy& Radiopharmaceuticals.2001,16:63-71.
[11]Pia Hartmann,Georg Plum.Immunological Defense Mechanisms inTuberculosis and MAC-Infection.Diagn Microbiol Infect Dis.1999,34:147-152.
[12]Padmini Salgame. Host innate and Thl responses and the bacterial factors that control Mycobacterium tuberculosis infection. Current Opinion in Immunology,2005,17:374-380.
[13]Csiszar A, Nagy G, Gergely P. Increased interferon-gamma, IL-10 and decreased IL-4 mRNA expression in peripheral blood mononuclear cells from patients with systemic lupus erythematosus. Clin Exp Immunol.2000,122:464-470.
[14]华树成,迟宝荣,李丹等.结核免疫及免疫治疗的进展.吉林大学学报(医学版).2005,31(6)：985-988.
[15]Siobha'n C. Cowleyl, Karen L. Elkins. CD4+T Cells Mediate IFN-γ-Independent Control of Mycobacterium tuberculosis Infection Both In Vitro and In Vivo. The Journal of Immunology.2003,171:4689-4699.
[16]Boehm, U., T. Klamp, M. Groot, J. C. Howard. Cellular responses to interferon-y. Annu. Rev. Immunology.1997,15:749.
[17]Rish K. Pai, Marilyn Convery, Thomas A. Hamilton, et al. Inhibition of IFN-y-Induced Class II Transactivator Expression by a 19-kDa Lipoprotein from Mycobacterium tuberculosis:A Potential Mechanism for Immune Evasion. The Journal of Immunology.2003,171:175-184.
[18]Sybille Thma-Uszynski, Steffen Stenger, Osamu Takeuchi, et al. Induction of Direct Antimicrobial Activity Through Mammalian Toll-like Receptors. Science.2001,291: 1544-1546.
[19]Sarah M. Fortune, Alejandra Solache, Alejandra Jaeger, et al. Mycobacterium tuberculosis Inhibits Macrophage Responses to IFN-y through Myeloid Differentiation Factor 88-Dependent and -Independent Mechanisms. The Journal of Immunology.2004,172: 6272-6280.
[20]Brandon M. Sullivan, Ousman Jobe, Vanja Lazarevic, et al. Increased Susceptibility of Mice Lacking T-bet to Infection with Mycobacterium tuberculosis Correlates with Increased IL-10 and Decreased IFN-y Production. The Journal of Immunology.2005,175:4593-4602.
[21]Muazzam Jacobs, Dieudonne'e Togbe, Cecile Fremond, et al. Tumor necrosis factor is critical to control tuberculosis infection. Microbes and Infection.2007,9:623-628.
[22]Andrea M. Cooper,Jeanne Magram, Jessica Ferrante, et al. Interleukin 12 (IL-12) Is Crucial to the Development of Protective Immunity in Mice Intravenously Infected With Mycobacterium tuberculosis. J. Exp. Med.1997,186:39-45.
[23]Dawn Noltl, JoAnne L. Flynn, Interleukin-12 Therapy Reduces the Number of Immune Cells and Pathology in Lungs of Mice Infected with Mycobacterium tuberculosis. Infection and Immunity.2004,72(5):2976-2988.
[24]Carl G. Feng, Dragana Jankovic, Marika Kullberg, et al. Maintenance of Pulmonary Th1 Effector Function in Chronic Tuberculosis Requires Persistent IL-12 Production. The Journal of Immunology.2005,174:4185-4192.
[25]Szabo, S.J., N.G. Jacobson, A.S. Dighe, U. Gubler, et al. Developmental commitment to the Th2 lineage by extinction of IL-12 signaling. Immunity.1995,2:665-675.
[26]CHANG-HWA SONG, HWA-JUNG KIM, JEONG-KYU PARK, et al. Depressed Interleukin-12 (IL-12), but not IL-18, Production in Response to a 30- or 32-Kilodalton Mycobacterial Antigen in Patients with Active Pulmonary Tuberculosis. Infection and Immunity.2000,68(8):4477-4484.
[27]U. Greinert, M. Ernst, M. Schlaak, P. Entzian, et al. Interleukin-12 as successful adjuvant in tuberculosis treatment. Eur Respir J.2001,17:1049-1051.
[28]Abebech Demissie, Markos Abebe, Abraham Aseffa, et al. Healthy Individuals That Control a Latent Infection with Mycobacterium tuberculosis Express High Levels of Thl Cytokines and the IL-4 Antagonist IL-482. The Journal of Immunology.2004,172: 6938-6943.
[29]Helen A.fletcher, Patrick Owiafe. David Jeffries, et al. Increased expression of mRNA encoding interleukin (IL)-4 and its splice variant IL-4d2 in cells from contacts of Mycobacterium tuberculosis, in the absence of in vitro stimulation. Immunology.2004. 112:669-673
[30]Graham A.W. Rook, Rogelio Hernandez-Pando, Keertan Dheda, et al. IL-4 in tuberculosis:implications for vaccine design. TRENDS in Immunology.2004,25 (9): 483-488.
[31]初乃惠,朱莉贞,叶志忠等.重组人白细胞介素-2辅助治疗复治肺结核近期疗效观察.中华结核和呼吸杂志,2003,26(9)：548-551.
[32]庄玉辉,李国利,王小宁等.rIL-2在小鼠体内抗结核菌能力的研究.上海免疫学杂志,1992,12(2)：76-77.
[33]Luiz. Eduardom. Bermudez, Paul Stevens, Peter Kolonoski, et al. Treatment of experimental disseminated Mycobacterium avium complex infection in mice with recombinant IL-2 and tumor necrosis factor. The journal of immunologist.1989, 143:2996-3000.
[34]John L. Johnson, Emmanuel Ssekasanvu, Alphonse Okwera, et al. Randomized Trial of Adjunctive Interleukin-2 in Adults with Pulmonary Tuberculosis. Am J Respir Crit Care Med.2003,168:185-191.
[35]Zahra Toossi, Mary Ellen Kleinhenz, Jerrold J. Ellner. Deffective interleukin 2 production and responsiveness in human pulmonary tuberculosis. Journal of Experimental Medicine.1986,163:1162-1172.
[36]H. Cai, D.H. Tian, Y.X. Zhu. Coadministration of Interleukin 2 plasmid DNA with combined DNA vaccines significantly enhances the protective efficacy against Mycobacterium tuberculosis. DNA and Cell Biology.2005,24(10):605-613.
[37]Long R & Gardam M. Tumour necrosis factor-alpha inhibitors and the reactivation of latent tuberculosis infection. CMAJ.2003,168(9):1153-1156.
[38]Saunders B, Tran S, Ruuls S. et al, Transmembrane TNF is suffi cient to initiate cell migration and granuloma formation and provide acute, but not long-term, control of Mycobacterium tuberculosis infection. J. Immunol.2005,174:4852-4859.
[39]Olleros, M. et al. Contribution of transmembrane tumor necrosis factor to host defense against Mycobacterium bovis bacillus Calmette-guerin and Mycobacterium tuberculosis infections. Am. J. Path.2005,166:1109-1120.
[40]Michael A Gardam, Edward C Keystone, Richard Menzies et al.Anti-tumour necrosis factor agents and tuberculosis risk:mechanisms of action and clinical management.Lancet Infect Dis,2003;3:148-55
[41]Muazzam Jacobs, Dieudonne'e Togbe, Cecile Fremond et al. Tumor necrosis factor is critical to control tuberculosis infection. Microbes and Infection,2007,9:623-628.
[42]Charles A. Dinarello. Anti-cytokine therapeutics and infections. Vaccine.2003,21: 24-34.
[43]Tomioka H,Namba K.Develepment of antituberculous drugs:current status and future prospects.Kekkaku.2006,81(12):753-74.
[44]S Stenger. Immunological control of tuberculosis:role of tumor necrosis factor and more. Ann Rheum Dis.2005,64:24-28.
[45]LP Ormerod. Tuberculosis and anti-TNF-atreatment. Thorax.2004,59:921.
[46]Suzuki K, Lee WJ, et al. Recombinant granulocyte-macrophage colony-stimulating factor (GM-CSF) or tumor necrosis factor-alpha (TNF-a) activate human alveolar macrophage to inhibit growth of Mycobacterium avium complex. Clin Exp Immunol.1994, 98:169-173.
[47]Gonzalez-Juarrero, et al., Disruption of granulocyte macrophage-colony stimulating factor production in the lungs severely affects the ability of mice to control Mycobacterium tuberculosis infection. J. Leukoc. Biol.2005,77:914-922.
[48]Szeliga, J., Daniel, D. S., et al. Granulocyte-macrophage colony stimulating factor-mediated innate responses in tuberculosis. Tuberculosis.2008,88:7-20.
[49]Zhang X, Divangahi M, Ngai P, et al. Intramuscular immunization with a monogenic plasmid DNA tuberculosis vaccine:Enhanced immunogenicity by electroporation and co-expression of GM-CSF transgene. Vaccine.2007,25:1342-1352.
[50]Ryan AA, Wozniak TM, Shklovskaya E, et al. Improved protection against disseminated tuberculosis by Mycobacterium bovis bacillus Calmette-Guerin secreting murine GM-CSF is associated with expansion and activation of APCs. J Immunol.2007,179:8418-8424.
[51]Nambiar JK, Ryan AA, Kong CU, et al. Modulation of pulmonary DC function by vaccine-encoded GM-CSF enhances protective immunity against Mycobacterium tuberculosis infection. European Journal of Immunology.2010,40:153-161.
[52]Jordan J D, Landac E M, Iyengar R. Signaling Networks:The Origins of Cellular Multasking. Cell.2000,103:193-200.
[53]Callard R, George A J, Srtark J. Cytokines, Chaos, and Complexity. Immunity.1999, 11:507-513.
[54]Ziv Frankenstein, Uri Alon, Irun R Cohen. The immune-body cytokine network defines a social architecture of cell interactions. Biology Direct.2006,1:32.
[55]Bartlett, J. G. Clostridium difficile:history of its role as an enteric pathogen and current state of knowledge about the organism. Clin. Infect. Dis.1994,18(4):265-272.
[56]Bustinza A, Solana MJ, Padilla B, et al. Nosocomial outbreak of Clostridium difficile-associated disease in a pediatric intensive care unit in Madrid. Infect Control Hosp Epidemiol.2009,30(2):199-201.
[57]Weiss K, Boisvert A, Chagnon M, et al. Multipronged intervention strategy to control an outbreak of Clostridium difficile infection (CDI) and its impact on the rates of CDI from 2002 to 2007. Infect Control Hosp Epidemiol.2009,30(2):156-62.
[58]Graf K, Cohrs A, Gastmeier P, An outbreak of Clostridium difficile-associated disease (CDAD) in a German university hospital. Eur J Clin Microbiol Infect Dis.2009,28(5):543-5.
[59]Loo VG, Poirier L, Miller MA, et al. A predominantly clonal multi-institutional outbreak of Clostridium difficile-associated diarrhea with high morbidity and mortality. N Engl J Med. 2005,353:2442-2449.
[60]Pepin J, Saheb N, Coulombe MA, et al. Emergence of fluoroquinolones as the predominant risk factor for Clostridium difficile-associated diarrhea:a cohort study during an epidemic in Quebec. Clin Infect Dis.2005,41:1254-1260.
[61]Oldfield EC 3rd. Clostridium difficile-associated diarrhea:resurgence with a vengeance. Rev Gastroenterol Disord.2006,6:79-96.
[62]Kim, J. et al. Epidemiological features of Clostridium difficile associated disease among inpatients at children's hospitals in the United States. Pediatrics.2008,122:1266-1270.
[63]Rouphael, N. G. et al. Clostridium difficile-associated diarrhea:an emerging threat to pregnant women. Am. J. Obstet. Gynecol.2008,198:635. el-e6.
[64]Huang H, Wu S, Wang M, et al. Clostridium difficile infections in a Shanghai hospital: antimicrobial resistance, toxin profiles and ribotype. Int Antimicrob Agents.2009,33(4): 339-342.
[65]Sayedy L, Kothari D, Richards RJ. Toxic megacolon associated Clostridium difficile colitis. World J Gastrointest Endosc.2010,2(8):293-7.
[66]Cheng, V. C., W. C. Yam, J. F. Chan, et al. Clostridium difficile ribotype 027 arrives in HongKong. Int. J. Antimicrob. Agengts.2009,34:492-493.
[67]Alexander Indra, Marion Blaschitz, Silvia Kernbichler, et al. Mechanisms behind variation in the Clostridium difficile 16S-23S rRNA intergenic spacer region. Journal of Medical Microbiology.2010,59:1317-1323.
[68]张维亚,张瑞.克林霉素副作用与艰难梭状芽孢杆菌的关系.中国实用医药.2009,4(30)：131-132.
[69]杨昭徐.PPI引起艰难梭状芽孢杆菌性腹泻的危险因素.中国医院用药评价与分析.2006,3：147-150.
[70]贾晋松,黄晓军,刘代红等.异基因造血干细胞移植过程中患者艰难梭菌感染与肠道微生态失调关系的临床研究.中国实验血液学杂志.2008,16(1)：135-139.
[71]Sandra Janezic, Maja Rupnik. Molecular typing methods for Clostridium difficile: pulsed-field gel electrophoresis and PCR ribotyping. Methods in Molecular Biology.2010, 646:55-65.
[72]Redelings MD, Sorvillo F, Mascola. Increase in Clostridium difficile related mortality rates, United States,1999-2004. Emerging Infectious Diseases.2007,13(9):1417-1419.
[73]J. Freeman, M. P. Bauer, S. D. Baines, et al. The changing epidemiology of clostridium difficile infections. Clinical Microbiology Reviews.2010,23(3):529-549.
[74]Richard A. Stabler, Lisa F. Dawson, Leslie T. H. Phua, et al. Comparative analysis of BI/NAP 1/027 hypervirulent strains reveals novel toxin B-encoding gene (tcdB) sequences. Journal of Medical Microbiology.2008,57:771-775.
[75]Richard A Stabler, Miao He, Lisa Dawson, et al. Comparative genome and phenotypic analysis of dseweez Clostridium difficile 027 strains provides insight into the evolution of a hypervirulent bacterium. Genome Biology.2009,10(9):102.1-102.15.
[76]Spigaglia P, Mastrantonio P. Analysis of macrolide-lincosamide-streptogramin B (MLS(B)) resistance determinant in strains of Clostridium difficile. icrob Drug Resist.2002, 8(1):45-53.
[77]L. Clifford McDonald, George E. Killgore, Angela Thompson, et al. An epidemic, toxin gene-variant strain of Clostridium difficile. The New England Journal of Medicine.2005, 353(23):2433-2441.
[78]Trevor D. Lawley, Simon Clare, Alan W. Walker, et al. Antibiotic treatment of clostridium difficile carrier mice triggers a supershedder state, spore-mediated transmission, and severe disease in immunocompromised hosts. Infection and Immunity.2009,77(9): 3661-3669.
[79]V. K. Viswanathan, M. J. Mallozzi, Gayatri Vedantam. Clostridium diffidicle infection an overview of the disease and its pathogenesis epidemiology and interventions. Gut Microbes. 2010,1(4):234-242.
[80]Yvonne Kleypas, Dennis McCubbin, Hlizabetb S. Curnow. The role of environmental cleaning in health care-associated infections. Critical Care Nursing Quarterly.2011,1(49): 11-17.
[81]Kato H, Kato N, Watanabe K. Relapses or reinfections:analysis of a case of Clostridium difficile-associated colitis by two typing systems. Curr Microbiol.1996,33(4):220-3.
[82]G. L. O'NEILL, M. H. BEAMAN, T. V. RILEY. Relapse versus reinfection with Clostridium diffcile. Epidemiol Infect.1991,107:627-635.
[83]She RC, Durrant RJ, Petti CA. Evaluation of enzyme immunoassays to detect Clostridium difficile toxin from anaerobic stool culture. Am J Clin Pathol.2009,131(1):81-4.
[84]C. Wenisch, B. Parschalk, M. Hasenbundl, A. M. Hirschl, et al. Comparison of vancomycin, teicoplanin, metronidazole, and fusidic acid for the treatment of clostridium difficile-associated diarrhea. Clinical Infectious Diseases.1996,22:813-8.
[85]Sullivan KM, Spooner LM. Fidaxomicin:a macrocyclic antibiotic for the management of Clostridium difficile infection. Ann Pharmacother.2010,44(2):352-9.
[86]Johnson S, Homann SR, Bettin KM, et al. Treatment of asymptomatic Clostridium difficile carriers (fecal excretors) with vancomycin or metronidazole, A randomized, placebo-controlled trial.1992,117(4):297-302.
[87]Alan R. Salkind, Clostridium difficile:an update for the primary care clinician. South Med J.2010,103(9):896-902.
[88]George Tannous, Guy Neff, Nyingi Kemmer. Therapeutic Success of Rifaximin for Clostridium difficile Infection Refractory to Metronidazole and Vancomycin. Gastroenterology.2010,4:404-409.
[89]Shah D, Dang MD, Hasbun R, et al. Clostridium difficile infection:update on emerging antibiotic treatment options and antibiotic resistance. Expert Rev Anti Infect Ther.2010,8(5): 555-64.
[90]Johnson AP. New antibiotics for selective treatment of gastrointestinal infection caused by Clostridium difficile. Expert Opin Ther Pat.2010,20(10):1389-99.
[91]Miller M. Fidaxomicin (OPT-80) for the treatment of Clostridium difficile infection. Expert Opin Pharmacother.2010,11(9):1569-78.
[92]Poxton IR. Fidaxomicin:a new macrocyclic, RNA polymerase-inhibiting antibiotic for the treatment of Clostridium difficile infections. Future Microbiology.2010,5(4):539-548.
[93]Tannock GW, Munro K, Taylor C, et al. A new macrocyclic antibiotic, fidaxomicin (OPT-80), causes less alteration to the bowel microbiota of clostridium difficile-infected patients than does vancomycin. Microbioligy.2010,156(11):3354-9.
[94]Thomas J. Louie, M.D., Mark A. Miller, M.D., Kathleen M. Mullane, et al. Fidaxomicin versus Vancomycin for Clostridium difficile Infection. The New England Journal of Medicine.2011,364:422-431.
[95]John G. Bartlett, Clostridium difficile:progress and challenge, Annals of the New York academy of sciences.2010,1213:62-69.
[96]Lsrael Lowy, Deborah C. Molrine, Brett A. Leav, et al. Treatment with monoclonal antibodies against Clostridium difficile toxins. The New England Journal of Medicine.2010, 362(3):197-205.
[97]Gregory J. Babcock, Teresa J. Broering, Hector J. Hernandez, et al. Human monoclonal antibodies directed against TcdA and TcdB prevent Clostridium difficile-induced mortality in hamsters. Infection and Immunity.2006,74(11):6339-6347.
[98]Tomoaki Saito, Soichiro Kimura, Kazuhiro Tateda, et al. Evidence of intravenous immunoglobulin as a critical supportive therapy against Clostridium difficile toxin-mediated lethality in mice. Journal of Antimicrobial Chemotherapy.2011,66(5):1096-1099.
[99]Hassoun A, Ibrahim F. Use of intravenous immunoglobulin for the treatment of severe Clostridium difficile colitis. Am J Geriatr Pharmacother.2007,5(1):48-51.
[100]Young KW, Munro IC, Taylor SL, et al. The safety of whey protein concentrate derived from the milk of cows immunized against Clostridium difficile, Regul Toxicol Pharmacol. 2007,47(3):317-26.
[101]Numan SC, Veldkamp P, Kuijper EJ. Clostridium difficile-associated diarrhoea:bovine anti-Clostridium difficile whey protein to help aid the prevention of relapses. Gut.2007, 56(6):888-9.
[102]Van Dissel JT, de Groot N, Hensgens CM. Bovine antibody-enriched whey to aid in the prevention of a relapse of Clostridium difficile-associated diarrhoea:preclinical and preliminary clinical data. J Med Microbiol.2005,54(2):197-205.
[103]KarlWeiss, Toxin-binding treatment for Clostridium difficile:a review including reports of studies with tolevamer, International Journal of Antimicrobial Agents.2009,33:4-7.
[104]Sonia S. Yoon, Lawrence J. Brandt. Treatment of Refractory/Recurrent C. difficile-associated Disease by Donated Stool Transplanted Via Colonoscopy A Case Series of 12 Patients. J Clin Gastroenterol.2010,44(8):562-566.
[105]Timothy A. Rubin, Charles E. Gessert, Johannes Aas, et al. Stool transplantation for older patients with Clostridium difficile infection. J Am Geriatr Soc.2009,57(12):2386-7.
[106]Russell G, Kaplan J, Ferraro M, et al. Fecal bacteriotherapy for relapsing Clostridium difficile infection in a child:a proposed treatment protocol. Pediatrics.2010,126(1):239-42.
[107]Von Eichel-Streiber, C. et al. Comparative sequence analysis of the Clostridium difficile toxins A and B, Mol. Gen. Genet.1992,233:260-268.
[108]Kelly CP, LaMont JT. Clostridium difficile---More difficult than ever. N Engl J Med. 2008,359:1932-40.
[109]Stuart H. Cohen, Yajarayma J. Tang, Joseph Silva. Analysis of the Pathogenicity Locus in Clostridium difficile Strains. The Journal of Infectious Diseases.2000,181:659-63.
[110]Maja Rupnik, Mark H. Wilcox, Dale N. Gerding. Clostridium difficile infection:new developments in epidemiology and pathogenesis. Nature Reviews Microbiology.2009, 7:526-536.
[111]I. Just·R. Gerhard. Large clostridial cytotoxins. Rev Physiol Biochem Pharmacol.2004, 152:23-47.
[112]Thomas Jank, Klaus Aktories. Structure and mode of action of clostridial glucosylating toxins:the ABCD model. Trends in Microbiology.2008,16(5):222-229.
[113]Markus Sauerborn, Petra Leukel, Christoph von Eichel-Streiber. The C-terminal ligand-binding domain of Clostridium difficile toxin A (TcdA) abrogates TcdA-specific binding to cells and prevents mouse lethality. FEMS Microbiology Letters.1997,155:45-54.
[114]Hofmann F, Busch C, Prepens U. Localization of the glucosyltransferase activity of Clostridium difficile toxin B to the N-terminal part of the holotoxin. J Biol Chem.1997, 272(17):11074-8.
[115]Thomas Jank, Torsten Giesemann, Klaus Aktories. Rho-glucosylating Clostridium difficile toxins A and B:new insights into structure and function. Clycobiology.2007,17(4): 15-22.
[116]Marinilce F. Santos, Shirley A. McCormack, Zhong Guo. Rho Proteins Play a Critical Role in Cell Migration during the Early Phase of Mucosal Restitution. Journal of Clinical Investigation.1997,100(1):216-255.
[117]Ismael Halabi-Cabezona, Johannes Huelsenbecka, Martin May. Prevention of the cytopathic e□ect induced by Clostridium di□cile Toxin B by active Racl. Federation of European Biochemical Societies.2008,582:3751-3756.
[118]Thomas Jank, Torsten Giesemann, Klaus Aktories. Clostridium difficile Glucosyltransferase Toxin B-essential Amino Acids for Substrate Binding. J Biol Chem.2007, 282 (48):35222-35231.
[119]Esteban Chaves-Olarte, Manfred Weidmann, Christoph von Eichel-Streiber. Toxins A and B from Clostridium difficile Differ with Respect to Enzymatic Potencies, Cellular Substrate Specificities and Surface Binding to Cultured Cells. J. Clin. Invest.1997, 100(7):1734-1741.
[120]Jessica Reineke, Stefan Tenzer, Maja Rupnik. Autocatalytic cleavage of Clostridium difficile toxin B. Nature.2007,446(7134):415-419.
[121]Martina Egerer, Torsten Giesemann, Christian Herrmann. Autocatalytic Processing of Clostridium difficile Toxin B Binding of Inositol Hexakisphosphate. J Biol Chem.2009, 284(6):3389-3395.
[122]Martina Egerer, Torsten Giesemann, Thomas Jank. Auto-catalytic Cleavage of Clostridium difficile Toxins A and B Depends on Cysteine Protease Activity. J Biol Chem. 2007,282(35):25314-25321.
[123]Isa Kreimeyer, Friederike Euler, Alexander Marckscheffel, et al. Autoproteolytic cleavage mediates cytotoxicity of Clostridium difficile toxin A. Naunyn-Schmied Arch Pharmacol.2011,383(3):253-262.
[124]Lehrer RI, Jung G, Ruchala P, et al. Human alpha-defensins inhibit hemolysis mediated by cholesterol-dependent cytolysins. Infect Immun.2009,77(9):4028-40.
[125]Giesemann T, Jank T, Gerhard R, et al. Cholesterol-dependent pore formation of Clostridium difficile toxin A. J Biol Chem.2006,281(16):10808-15.
[126]Selda Genisyuerek, Panagiotis Papatheodorou, Gregor Guttenberg. Structural Determinants for Membrane Insertion, Pore Formation and Translocation of Clostridium difficile Toxin B. Molecular Microbiology.2011,79(6):1643-1654.
[127]Jason G. S. Ho, Antonio Greco, Maja Rupnik, et al. Crystal structure of receptor-binding C-terminal repeats from Clostridium difficile toxin A. PNAS.2005,102 (51):18373-18378.
[128]Cornelia Frisch, Ralf Gerhard, Klaus Aktories. The complete receptor-binding domain of Clostridium difficile toxin A is required for endocytosis. Biochemical and Biophysical Research Communications.2003,300:706-711.
[129]Tanis Dingle, Stefanie Wee, George L Mulvey. Functional properties of the carboxy-terminal host cell-binding domains of the two toxins, TcdA and TcdB, expressed by Clostridium difficile. Glycobiology.2008,18(9):698-706.
[130]Xi Na, Ho Kim,1 Mary P. Moyer, et al. gp96 Is a Human Colonocyte Plasma Membrane Binding Protein for Clostridium difficile Toxin A. Infection and Immunity.2008, 76(7):2862-2871.
[131]M. K. KEEL, J. G. SONGER. The Distribution and Density of Clostridium difficile Toxin Receptors on the Intestinal Mucosa of Neonatal Pigs. Vet Pathol.2007,44:814-822.
[132]Maen Qa'Dan, Matthew Ramsey, Jeremy Daniel, et al. Clostridium difficiletoxin B activates dual caspase-dependent and caspase-independent apoptosis in intoxicated cells. Cellular Microbiology.2002,4(7):425-434.
[133]D. HE, S. J. HAGEN, C. POTHOULAKIS, et al. Clostridium difficile Toxin A Causes Early Damage to Mitochondria in Cultured Cells. gastroenterology.2000,119:139-150.
[134]Ho Kim, Sang Hoon Rhee, Efi Kokkotou, et al. Clostridium difficile Toxin A Regulates Inducible Cyclooxygenase-2 and Prostaglandin E2 Synthesis in Colonocytes via Reactive Oxygen Species and Activation of p38 MAPK. J. Bio.Chem.2005,280(22):21237-21245.
[135]Dan He, Stavros Sougioultzis, Susan Hagen, et al. Clostridium difficile Toxin A Triggers Human Colonocyte IL-8 Release via Mitochondrial Oxygen Radical Generation. gastroenterology.2002,122:1048-1057.
[136]Jin Young Lee, Hyunah Kim, Mi Yeon Cha, et al. Clostridium difficile toxin A promotes dendritic cell maturation and chemokine CXCL2 expression through p38, IKK, and the NF-κB signaling pathway. J Mol Med.2009,87:169-180.
[137]Paola Matarrese, Loredana Falzano, Alessia Fabbri, et al. Clostridium difficile Toxin B Causes Apoptosis in Epithelial Cells by Thrilling Mitochondria Involvement of ATP-Sensitive mitochondrial Potassium Channels. J Biol Chem.2007,282(12):9029-9041.
[138]Pothoulakis C., Lamont J.T., et al. Microbes and microbial toxins:paradigms for microbial-mucosal interactions Ⅱ. The integrated response of the intestine to Clostridium difficile toxins. Am J Physiol Gastrointest Liver Physiol.2001,280:G178-183.
[139]Kelly C.P., Becker S., Linevsky J.K., et al. Neutrophil recruitment in Clostridium difficile toxin A enteritis in the rabbit. J. Clin. Invest.1994,93:1257-1265.
[140]Lyerly D.M., Lockwood D.E., Richardson S.H., et al. Biological activities of toxins A and B of Clostridium difficile. Infect. Immun.1982,35:1147-1150.
[141]Lyerly D.M., Saum K.E., MacDonald D.K., et al. Effects of Clostridium difficile toxins given intragastrically to animals. Infect. Immun.1985,47:349-352.
[142]Mitchell T.J., Ketley J.M., Haslam S.C., et al. Effect of toxin A and B of Clostridium difficile on rabbit ileum and colon. Gut.1986,27:78-85.
[143]Lima A.A., Lyerly D.M., Wilkins T.D.,et al. Effects of Clostridium difficile toxins A and B in rabbit small and large intestine in vivo and on cultured cells in vitro. Infect. Immun.1988, 56:582-588.
[144]Bongaerts, Lyerly. Role of toxins A and B in the pathogenesis of Clostridium difficile disease. Infect. Immun.1994,17:1-12.
[145]Ketley J.M., Mitchell T.J., Candy D.C., et al. The effects of Clostridium difficile crude toxins and toxin A on ileal and colonic loops in immune and non-immune rabbits. J. Med. Microbiol.1987,24:41-52.
[146]Riegler M., Sedivy R., Pothoulakis C., et al. Clostridium difficile toxin B is more potent than toxin A in damaging human colonic epithelium in vitro. J. Clin. Invest.1995,95: 2004-2011.
[147]Savidge T.C., Pan W.H., Newman P, et al. Clostridium difficile toxin B is an inflammatory enterotoxin in human intestine. Gastroenterology.2003,125: 413-420.
[148]Borriello S.P., Wren B.W., Hyde S., et al. Molecular, immunological, and biological characterization of a toxin A-negative, toxin B-positive strain of Clostridium difficile. Infect. Immun.1992,60:4192-4199.
[149]Lyras D., O'Connor J.R., Howarth P.M., et al. Toxin B is essential for virulence of Clostridium difficile. Nature.2009,458:1176-1179.
[150]Geric, B., Carman, Rupnik, et al. Binary toxin-producing, large clostridial toxin-negative Clostridium difficile strains are enterotoxic but do not cause disease in hamsters. J. Infect. Dis.2006,193:1143-1150.
[151]Schwan, C. Stecher, B.Tzivelekidis, et al. Clostridium difficile toxin CDT induces formation of microtubule-based protrusions and increases adherence of bacteria. PLoS Pathog. 2009,5:e1000626.
[152]Qiu B., Pothoulakis C., Castagliuolo I., et al. Participation of reactive oxygen metabolites in Clostridium difficile toxin A-induced enteritis in rats. Am. J. Physiol.1999, 276:G485-G490.
[153]Xingmin Sun, Tor Savidge, Hanping Feng. The Enterotoxcity of Clostridium difficile. Toxins.2010,2:1848-1880.
[154]Qiu B., Pothoulakis C., Castagliuolo I., et al. Nitric oxide inhibits rat intestinal secretion by Clostridium difficile toxin A but not Vibrio cholerae enterotoxin. Gastroenterology.1996, 111:409-418.
[155]Savidge TC, Urvil P, Oezguen N, et al. Host S-nitrosylation inhibits clostridial small molecule-activated glucosylating toxins. Nature Medicine.2011,17(9):1136-41.
[156]Meyer G.K., Neetz A., Brandes, G, et al. Clostridium difficile toxins A and B directly stimulate human mast cells. Infect. Immun.2007,75:3868-3876.
[157]Lima A.A., Nascimento N.R., Fang G.D., et al. Role of phospholipase A2 and tyrosine kinase in Clostridium difficile toxin A-induced disruption of epithelial integrity, histologic inflammatory damage and intestinal secretion.J. Appl. Toxicol.2008,28:849-857.
[158]Hayashi H., Szaszi K., Coady-Osberg N., et al. Inhibition and redistribution of NHE3, the apical Na+/H+exchanger, by Clostridium difficile toxin B. J. Gen. Physiol.2004,123: 491-504.
[159]Alcantare C.S., Jin X.H., Brito G.A., et al. Angiotensin II subtype 1 recptor blockade inhibits Clostridium difficile toxin A-induced intestinal secretion in a rabbit model. J. Infect. Dis.2005,191:2090-2096.
[160]Na X., Zhao D., Koon H.W., et al. Clostridium difficile toxin B activates the EGF receptor and the ERK/MAP kinase pathway in human colonocytes. Gastroenterology.2005, 128:1002-1011.
[161]Calabi E., Fairweather N. Patterns of sequenc conservation in the S-layer proteins and related sequences in Clostridium difficile. J. Bacteriol.002,184:33886-33897.
[162]Drudy D. et al. Human antibody response to surface layer proteins in Clostridium difficile infection. FEMS Immunol. Med. Microbiol.2004,41:237-242.
[163]Wright A. et al. Proteomic analysis of cell surface proteins from Clostridium difficile. Proteomics.2005,5:2443-2452.
[164]Merrigan MM, Gerding DN, Vedantam G. Hypervirulent Clostridium difficile strains have altered protein expression and host-cell adherence. Eighth Biennial Conference of Anaerobe Society of America PI-12 (Boise, Idaho).2006.
[165]Pothoulakis C., Sullivan R., Melnick D.A., et al. Clostridium difficile toxin A stimulates intracellular calcium release and chemotactic response in human granulocytes. Am. J. Pathol. 1988,81:1741-1745.
[166]Ishida Y., Maegawa T., Kondo T., et al. Essential involvement of IFN-gamma in Clostridium difficile toxin A-induced enteritis. J. Immunol.2004,172:3018-3025.
[167]Krivan H.C., Clark G.F., Smith D.F., et al. Cell surface binding site for Clostridium difficile enterotoxin:evidence for a glycoconjugate containing the sequence Gal alpha 1-3Gal beta 1-4GlcNAc. Infect. Immun.1986,53:573-581.
[168]Pothoulakis C., Castagliuolo I., LaMont J.T. Nerves and Intestinal Mast Cells Modulate Responses to Enterotoxins. News Physiol. Sci.1998,13:58-63.
[169]Wershil B.K., Castagliuolo I., Pothoulakis C. Direct evidence of mast cell involvement in Clostridium difficile toxin A-induced enteritis in mice. Gastroenterology.1998,114: 956-964.
[170]Steinman R.M. The dendritic cell system and its role in immunogenicity. Annu. Rev. Immunol.1991,9:271-296.
[171]Lee J.Y., Kim H., Cha M.Y., et al. Clostridium difficile toxin A promotes dendritic cell maturation and chemokine CXCL2 expression through p38, IKK, and the NF-kappaB signaling pathway. J. Mol. Med.2009,87:169-180.
[172]Souza M.H., Melo-Filho A.A., Rocha M.F., et al. The involvement of macrophage-derived tumour necrosis factor and lipoxygenase products on the neutrophil recruitment induced by Clostridium difficile toxin B. Immunology.1997,91:281-288.
[173]Linevsky JK, Pothoulakis C, Keates S, et al. IL-8 release and neutrophil activation by Clostridium difficile toxin-exposed human monocytes. Am J Physiol.1997,273:1333-1340.
[174]Ng J., Hirota S.A., Gross O., et al. Clostridium difficile toxin-induced inflammation and intestinal injury are mediated by the inflammasome. Gastroenterology.2010,139(2):542-552.
[175]Rocha M.F., Maia M.E., Bezerra L.R., et al. Clostridium difficile toxin A induces the release of neutrophil chemotactic factors from rat peritoneal macrophages:role of interleukin-lbeta, tumor necrosis factor alpha, and leukotrienes. Infect. Immun.1997,65: 2740-2746.
[176]Flegel W.A., Muller F., Daubener W., et al. Cytokine response by human monocytes to Clostridium difficile toxin A and toxin B. Infect. Immun.1991,59:3659-3666.
[177]Abrams G.D., Allo M., Rifkin G.D., et al. Mucosal damage mediated by clostridial toxin in experimental clindamycin-associated colitis. Gut.1980,21,493-499.
[178]Czuprynski C.J., Johnson W.J., Balish E., et al. Pseudomembranous colitis in Clostridium difficile-monoassociated rats. Infect. Immun.1983,39:1368-1376.
[179]Fekety R., Silva J., Toshniwal R., et al. Antibiotic-associated colitis:effects of antibiotics on Clostridium difficile and the disease in hamsters. Rev. Infect. Dis.1979,1: 386-397.
[180]Knoop F.C. Clindamycin-associated enterocolitis in guinea pigs:evidence for a bacterial toxin. Infect. Immun.1979,23:31-33.
[181]Chang T.W., Bartlett J.G., Gorbach S.L., et al. Clindamycin-induced enterocolitis in hamsters as a model of pseudomembranous colitis in patients. Infect. Immun.1978,20: 526-529.
[182]Cormier G., Dubos F., Raibaud P. Modulation of cytotoxin production by Clostridium difficile in the intestinal tracts of gnotobiotic mice inoculated with various human intestinal bacteria. Appl. Environ. Microbiol.1985,49:250-252.
[183]Lusk R.H., Fekety R., Silva J., Browne R.A., et al. Clindamycin-induced enterocolitis in hamsters. J. Infect. Dis.1978,137:464-475.
[184]Price A.B., Larson H.E., Crow J. Morphology of experimental antibiotic-associated enterocolitis in the hamster:a model for human pseudomembranous colitis and antibiotic-associated diarrhoea. Gut 1979,20:467-475.
[185]Rehg J.E., Pakes S.P. Implication of Clostridium difficile and Clostridium perfringens iota toxins in experimental lincomycin-associated colitis of rabbits. Lab. Anim. Sci.1982,32: 253-257.
[186]Sugiyama T., Mukai M., Yamashita R., et al. Experimental models of Clostridium difficile enterocolitis in gnotobiotic mice. Prog. Clin. Biol. Res.1985,181:203-206.
[187]Chen X., Katchar K., Goldsmith J.D., et al. A mouse model of Clostridium difficile-associated disease. Gastroenterology 2008,135:1984-1992.
[188]Steele J., Feng H., Parry N., et al. Piglet models of acute or chronic Clostridium difficile illness. J. Infect. Dis.2010,201:428-434.
[189]Hamm E.E., Voth D.E., Ballard J.D. Identification of Clostridium difficile toxin B cardiotoxicity using a zebrafish embryo model of intoxication. Proc. Natl. Acad. Sci.2006, 103:14176-14181.
[190]Rolfe R.D., Iaconis J.P. Intestinal colonization of infant hamsters with Clostridium difficile. Infect. Immun.1983,42:480-486.
[191]M. K. Keel, J. G. Songer. The distribution and density of Clostridium difficile toxin receptors on the intestinal mucosa of neonatal pigs. Vet Pathol.2007,44:814-822.
[192]Hippenstiel S., Schmeck B., N'Guessan P.D., et al. Rho protein inactivation induced apoptosis of cultured human endothelial cells. Am. J. Physiol. Lung Cell. Mol. Physiol.2002, 283, L830-L838.
[193]Sarah A. Kuehnel, Stephen T. Cartmanl, John T. Heap, et al. The role of toxin A and toxin B in Clostridium difficile infection. Nature.2010,467(7316):711-713.
[194]Tim Du MScl, Michelle J Alfa. Translocation of Clostridium difficiletoxin B across polarized Caco-2 cell monolayers is enhanced by toxin A. Can J Infect Dis.2004,15(2): 83-88.
[195]P. A. Sutton, S. Li, J. Webb, et al. Essential role of toxin A in C. difficile 027 and reference strain supernatant-mediated disruption of Caco-2 intestinal epithelial barrier function. Clinical and Experimental Immunology.2008,153:439-447.
[196]Arnaud L, Gorochov G., Charlotte F, Lvovschi V, Parizot C, et al. Systemic perturbation of cytokine and chemokine networks in Erdheim-Chester disease:a single-center series of 37 patients. Blood.2011,117:2783-2790.
[197]Katsikis PD, Mueller YM, Villinger F. The Cytokine Network of Acute HIV Infection: A Promising Target for Vaccines and Therapy to Reduce Viral Set-Point? PLoS Pathog.2011, 7:e1002055.
[198]Cai H, Yu DH, Tian X, Zhu YX. Coadministration of interleukin 2 plasmid DNA with combined DNA vaccines significantly enhances the protective efficacy against Mycobacterium tuberculosis. DNA Cell Biol.2005,24:605-613.
[199]Nambiar JK, Ryan AA, Kong CU, et al. Modulation of pulmonary DC function by vaccine-encoded GM-CSF enhances protective immunity against Mycobacterium tuberculosis infection. Eur J Immunol.2010,40:153-161.
[200]Zhuang Y, Li G, Zhang X, et al. rIL-2 zai xiao shu ti nei kang jie he neng li de yan jiu. Current Immunology.1992,12:3.
[201]Szeliga J, Daniel DS, Yang CH, et al. Granulocyte-macrophage colony stimulating factor-mediated innate responses in tuberculosis. Tuberculosis.2008,88:7-20
[202]Toubaji A, Hill S, Terabe M, et al. The combination of GM-CSF and IL-2 as local adjuvant shows synergy in enhancing peptide vaccines and provides long term tumor protection. Vaccine.2007,25:5882-5891.
[203]Ahlers JD, Dunlop N, Alling DW, et al. Cytokine-in-adjuvant steering of the immune response phenotype to HIV-1 vaccine constructs:granulocyte-macrophage colony-stimulating factor and TNF-alpha synergize with IL-12 to enhance induction of cytotoxic T lymphocytes. J Immunol.1997,158:3947-3958.
[204]Orlova MO, Majorov KB, Lyadova IV, et al. Constitutive differences in gene expression profiles parallel genetic patterns of susceptibility to tuberculosis in mice. Infect Immun.2006, 74:3668-3672.
[205]Ota H, Kumagai S, Morinobu A, et al. Enhanced production of transforming growth factor-beta (TGF-beta) during autologous mixed lymphocyte reaction of systemic sclerosis patients. Clin Exp Immunol.1995,100:99-103.
[206]Dheda K, Chang JS, Breen RA, et al. In vivo and in vitro studies of a novel cytokine, interleukin 4delta2, in pulmonary tuberculosis. Am J Respir Crit Care Med.2005,172: 501-508.
[207]Rial D. Rolfe. Binding kinetics of Clostridium difficile toxins A and B to intestinal brush border membranes from infant and adult hamsters. Infection and Immunity.1991,59(4): 1223-1230.
[208]Barroso LA, Moncrief JS, Lyerly DM, Wilkins TD. Mutagenesis of the Clostridium difficile toxin B and effect on cytotixic activity. Microb Pathog.1994,16(4):297-303.
[209]David Albesa-Jove, Thomas Bertrand, Elisabeth P. Carpenter. Four Distinct Structural Domains in Clostridium difficile Toxin B Visualized Using SAXS. J. Mol. Biol.2010,396: 1260-1270.
[210]Rory N. Pruitt, Melissa G. Chambers, Kenneth K.-S. Ng, et al. Structural organization of the functional domains of Clostridium difficile toxins A and B. PNAS.2010,107(30): 13467-72.
[211]von Eichel-Streiber C, Boquet P, Sauerborn M, et al. Large clostridial cytotoxins-a family of glycosyltransferases modifying small GTP-binding proteins. Trends Microbiol. 1996,4:375-382.
[212]Xingmin Sun, Xiangyun He, Saul Tzipori, et al. Essential role of the glucosyltransferase activity in Clostridium difficile toxin-induced secretion of TNF-a by macrophages. Microbial Pathogenesis.2009,46(6):298-305.
[213]Torsten Giesemann, Gregor Guttenberg, Klaus Aktories, Human a-Defensins Inhibit Clostridium difficile Toxin B. Gastroenterology.2008,134:2049-2058.
[214]Daniel M. Musher, Saima Aslam, Nancy Logan, et al. Relatively poor outcome after treatment of Clostridium difficile colitis with metronidazole. Clin Infect Dis.2005,40(11): 1586-1.
[215]John G. Bartlett. Clostridium difficile:progress and challenges, Annals of the New York Academy of Science.2010,1213:62-69.
[216]Johannes Huelsenbeck, Stefanie Dreger, Ralf Gerhard, et al. Difference in the Cytotoxic Effects of Toxin B from Clostridium difficile Strain VPI 10463 and Toxin B from Variant Clostridium difficile Strain 1470. Inf. Immu.2007,75(2):801-809.
[217]Antonio Greco, Jason G S Ho, Shuang-Jun Lin, et al. Carbohydrate recognition by Clostridium difficile toxin A. Nature Structural & Biology.2006,13(5):460-461.
[218]Chiou-Yueh Yeh, Chun-Nan Lin, Chuan-Fa Chang, et al. C-Terminal Repeats of Clostridium difficile Toxin A Induce Production of Chemokine and Adhesion Molecules in Endothelial Cells and Promote Migration of Leukocytes, Infection and Immunity.2008,76(3): 1170-1178.
[219]Martin Riegler, Roland Sedivy, Charalabos Pothoulakis, Gerhard Hamilton, et al. Clostridium difficile toxin B is more potent than toxin A in damaging human colonic epithelium in vitro. Journal of Clinical Investigation.1995,95:2004-2011.
[220]Jennifer Steele, Kevin Chen, Xingmin Sun, et al. Systemic Dissemination of Clostridium difficile Toxins A and B is Associated with Severe, Fatal Disease in Animal Models, The Journal of Infectious Diseases.2011 (accepted).
[2]彭卫生,王英年,肖成志等.新编结核病学.第二版.北京：中国医药科技出版社,2003.
[3]黄晓燕,严杰,李淑萍.结核分枝杆菌感染细胞的信号传导通路研究进展,中国生物制品学杂志,2006,19(3)：323-325.
[4]肖和平.结核病防治新进展.上海：复旦大学出版社,2004.
[5]谢惠安,肖成志,宋礼章.结核病化学疗法与化疗实施.北京：化学工业出版社,2005.
[6]Current development and future prospects in chemotherapy of tuberculosis
[7]王莉.浅谈细胞因子的网络性.河北省科学院学报,2002,19：92-97.
[8]唐神结,肖和平.细胞因子网络和结核病.中华结核和呼吸杂志,2001,24(9)：571-574.
[9]Pellegrini P, Contasta I, Berghella AM, et al. The TH1 and TH2 cytokine network in healtyh subjects:suggestions for experimental studies to create prognostic and diagnostic indices for biotherapeutic treatments[J]. Cancer Biotherapy & Radiopharmaceuticals.2000, 15:267-278.
[10]Contasta I, Pellegrini P, Berghella AM, et al. Cell cycle control in cellular homeostasis during the immune response interactions betweenTH1, TH2 cytokines, and Bcl2 and p53 molecules[J]. Cancer Biotherapy& Radiopharmaceuticals.2001,16:63-71.
[11]Pia Hartmann,Georg Plum.Immunological Defense Mechanisms inTuberculosis and MAC-Infection.Diagn Microbiol Infect Dis.1999,34:147-152.
[12]Padmini Salgame. Host innate and Thl responses and the bacterial factors that control Mycobacterium tuberculosis infection. Current Opinion in Immunology,2005,17:374-380.
[13]Csiszar A, Nagy G, Gergely P. Increased interferon-gamma, IL-10 and decreased IL-4 mRNA expression in peripheral blood mononuclear cells from patients with systemic lupus erythematosus. Clin Exp Immunol.2000,122:464-470.
[14]华树成,迟宝荣,李丹等.结核免疫及免疫治疗的进展.吉林大学学报(医学版).2005,31(6)：985-988.
[15]Siobha'n C. Cowleyl, Karen L. Elkins. CD4+T Cells Mediate IFN-γ-Independent Control of Mycobacterium tuberculosis Infection Both In Vitro and In Vivo. The Journal of Immunology.2003,171:4689-4699.
[16]Boehm, U., T. Klamp, M. Groot, J. C. Howard. Cellular responses to interferon-y. Annu. Rev. Immunology.1997,15:749.
[17]Rish K. Pai, Marilyn Convery, Thomas A. Hamilton, et al. Inhibition of IFN-y-Induced Class II Transactivator Expression by a 19-kDa Lipoprotein from Mycobacterium tuberculosis:A Potential Mechanism for Immune Evasion. The Journal of Immunology.2003,171:175-184.
[18]Sybille Thma-Uszynski, Steffen Stenger, Osamu Takeuchi, et al. Induction of Direct Antimicrobial Activity Through Mammalian Toll-like Receptors. Science.2001,291: 1544-1546.
[19]Sarah M. Fortune, Alejandra Solache, Alejandra Jaeger, et al. Mycobacterium tuberculosis Inhibits Macrophage Responses to IFN-y through Myeloid Differentiation Factor 88-Dependent and -Independent Mechanisms. The Journal of Immunology.2004,172: 6272-6280.
[20]Brandon M. Sullivan, Ousman Jobe, Vanja Lazarevic, et al. Increased Susceptibility of Mice Lacking T-bet to Infection with Mycobacterium tuberculosis Correlates with Increased IL-10 and Decreased IFN-y Production. The Journal of Immunology.2005,175:4593-4602.
[21]Muazzam Jacobs, Dieudonne'e Togbe, Cecile Fremond, et al. Tumor necrosis factor is critical to control tuberculosis infection. Microbes and Infection.2007,9:623-628.
[22]Andrea M. Cooper,Jeanne Magram, Jessica Ferrante, et al. Interleukin 12 (IL-12) Is Crucial to the Development of Protective Immunity in Mice Intravenously Infected With Mycobacterium tuberculosis. J. Exp. Med.1997,186:39-45.
[23]Dawn Noltl, JoAnne L. Flynn, Interleukin-12 Therapy Reduces the Number of Immune Cells and Pathology in Lungs of Mice Infected with Mycobacterium tuberculosis. Infection and Immunity.2004,72(5):2976-2988.
[24]Carl G. Feng, Dragana Jankovic, Marika Kullberg, et al. Maintenance of Pulmonary Th1 Effector Function in Chronic Tuberculosis Requires Persistent IL-12 Production. The Journal of Immunology.2005,174:4185-4192.
[25]Szabo, S.J., N.G. Jacobson, A.S. Dighe, U. Gubler, et al. Developmental commitment to the Th2 lineage by extinction of IL-12 signaling. Immunity.1995,2:665-675.
[26]CHANG-HWA SONG, HWA-JUNG KIM, JEONG-KYU PARK, et al. Depressed Interleukin-12 (IL-12), but not IL-18, Production in Response to a 30- or 32-Kilodalton Mycobacterial Antigen in Patients with Active Pulmonary Tuberculosis. Infection and Immunity.2000,68(8):4477-4484.
[27]U. Greinert, M. Ernst, M. Schlaak, P. Entzian, et al. Interleukin-12 as successful adjuvant in tuberculosis treatment. Eur Respir J.2001,17:1049-1051.
[28]Abebech Demissie, Markos Abebe, Abraham Aseffa, et al. Healthy Individuals That Control a Latent Infection with Mycobacterium tuberculosis Express High Levels of Thl Cytokines and the IL-4 Antagonist IL-482. The Journal of Immunology.2004,172: 6938-6943.
[29]Helen A.fletcher, Patrick Owiafe. David Jeffries, et al. Increased expression of mRNA encoding interleukin (IL)-4 and its splice variant IL-4d2 in cells from contacts of Mycobacterium tuberculosis, in the absence of in vitro stimulation. Immunology.2004. 112:669-673
[30]Graham A.W. Rook, Rogelio Hernandez-Pando, Keertan Dheda, et al. IL-4 in tuberculosis:implications for vaccine design. TRENDS in Immunology.2004,25 (9): 483-488.
[31]初乃惠,朱莉贞,叶志忠等.重组人白细胞介素-2辅助治疗复治肺结核近期疗效观察.中华结核和呼吸杂志,2003,26(9)：548-551.
[32]庄玉辉,李国利,王小宁等.rIL-2在小鼠体内抗结核菌能力的研究.上海免疫学杂志,1992,12(2)：76-77.
[33]Luiz. Eduardom. Bermudez, Paul Stevens, Peter Kolonoski, et al. Treatment of experimental disseminated Mycobacterium avium complex infection in mice with recombinant IL-2 and tumor necrosis factor. The journal of immunologist.1989, 143:2996-3000.
[34]John L. Johnson, Emmanuel Ssekasanvu, Alphonse Okwera, et al. Randomized Trial of Adjunctive Interleukin-2 in Adults with Pulmonary Tuberculosis. Am J Respir Crit Care Med.2003,168:185-191.
[35]Zahra Toossi, Mary Ellen Kleinhenz, Jerrold J. Ellner. Deffective interleukin 2 production and responsiveness in human pulmonary tuberculosis. Journal of Experimental Medicine.1986,163:1162-1172.
[36]H. Cai, D.H. Tian, Y.X. Zhu. Coadministration of Interleukin 2 plasmid DNA with combined DNA vaccines significantly enhances the protective efficacy against Mycobacterium tuberculosis. DNA and Cell Biology.2005,24(10):605-613.
[37]Long R & Gardam M. Tumour necrosis factor-alpha inhibitors and the reactivation of latent tuberculosis infection. CMAJ.2003,168(9):1153-1156.
[38]Saunders B, Tran S, Ruuls S. et al, Transmembrane TNF is suffi cient to initiate cell migration and granuloma formation and provide acute, but not long-term, control of Mycobacterium tuberculosis infection. J. Immunol.2005,174:4852-4859.
[39]Olleros, M. et al. Contribution of transmembrane tumor necrosis factor to host defense against Mycobacterium bovis bacillus Calmette-guerin and Mycobacterium tuberculosis infections. Am. J. Path.2005,166:1109-1120.
[40]Michael A Gardam, Edward C Keystone, Richard Menzies et al.Anti-tumour necrosis factor agents and tuberculosis risk:mechanisms of action and clinical management.Lancet Infect Dis,2003;3:148-55
[41]Muazzam Jacobs, Dieudonne'e Togbe, Cecile Fremond et al. Tumor necrosis factor is critical to control tuberculosis infection. Microbes and Infection,2007,9:623-628.
[42]Charles A. Dinarello. Anti-cytokine therapeutics and infections. Vaccine.2003,21: 24-34.
[43]Tomioka H,Namba K.Develepment of antituberculous drugs:current status and future prospects.Kekkaku.2006,81(12):753-74.
[44]S Stenger. Immunological control of tuberculosis:role of tumor necrosis factor and more. Ann Rheum Dis.2005,64:24-28.
[45]LP Ormerod. Tuberculosis and anti-TNF-atreatment. Thorax.2004,59:921.
[46]Suzuki K, Lee WJ, et al. Recombinant granulocyte-macrophage colony-stimulating factor (GM-CSF) or tumor necrosis factor-alpha (TNF-a) activate human alveolar macrophage to inhibit growth of Mycobacterium avium complex. Clin Exp Immunol.1994, 98:169-173.
[47]Gonzalez-Juarrero, et al., Disruption of granulocyte macrophage-colony stimulating factor production in the lungs severely affects the ability of mice to control Mycobacterium tuberculosis infection. J. Leukoc. Biol.2005,77:914-922.
[48]Szeliga, J., Daniel, D. S., et al. Granulocyte-macrophage colony stimulating factor-mediated innate responses in tuberculosis. Tuberculosis.2008,88:7-20.
[49]Zhang X, Divangahi M, Ngai P, et al. Intramuscular immunization with a monogenic plasmid DNA tuberculosis vaccine:Enhanced immunogenicity by electroporation and co-expression of GM-CSF transgene. Vaccine.2007,25:1342-1352.
[50]Ryan AA, Wozniak TM, Shklovskaya E, et al. Improved protection against disseminated tuberculosis by Mycobacterium bovis bacillus Calmette-Guerin secreting murine GM-CSF is associated with expansion and activation of APCs. J Immunol.2007,179:8418-8424.
[51]Nambiar JK, Ryan AA, Kong CU, et al. Modulation of pulmonary DC function by vaccine-encoded GM-CSF enhances protective immunity against Mycobacterium tuberculosis infection. European Journal of Immunology.2010,40:153-161.
[52]Jordan J D, Landac E M, Iyengar R. Signaling Networks:The Origins of Cellular Multasking. Cell.2000,103:193-200.
[53]Callard R, George A J, Srtark J. Cytokines, Chaos, and Complexity. Immunity.1999, 11:507-513.
[54]Ziv Frankenstein, Uri Alon, Irun R Cohen. The immune-body cytokine network defines a social architecture of cell interactions. Biology Direct.2006,1:32.
[55]Bartlett, J. G. Clostridium difficile:history of its role as an enteric pathogen and current state of knowledge about the organism. Clin. Infect. Dis.1994,18(4):265-272.
[56]Bustinza A, Solana MJ, Padilla B, et al. Nosocomial outbreak of Clostridium difficile-associated disease in a pediatric intensive care unit in Madrid. Infect Control Hosp Epidemiol.2009,30(2):199-201.
[57]Weiss K, Boisvert A, Chagnon M, et al. Multipronged intervention strategy to control an outbreak of Clostridium difficile infection (CDI) and its impact on the rates of CDI from 2002 to 2007. Infect Control Hosp Epidemiol.2009,30(2):156-62.
[58]Graf K, Cohrs A, Gastmeier P, An outbreak of Clostridium difficile-associated disease (CDAD) in a German university hospital. Eur J Clin Microbiol Infect Dis.2009,28(5):543-5.
[59]Loo VG, Poirier L, Miller MA, et al. A predominantly clonal multi-institutional outbreak of Clostridium difficile-associated diarrhea with high morbidity and mortality. N Engl J Med. 2005,353:2442-2449.
[60]Pepin J, Saheb N, Coulombe MA, et al. Emergence of fluoroquinolones as the predominant risk factor for Clostridium difficile-associated diarrhea:a cohort study during an epidemic in Quebec. Clin Infect Dis.2005,41:1254-1260.
[61]Oldfield EC 3rd. Clostridium difficile-associated diarrhea:resurgence with a vengeance. Rev Gastroenterol Disord.2006,6:79-96.
[62]Kim, J. et al. Epidemiological features of Clostridium difficile associated disease among inpatients at children's hospitals in the United States. Pediatrics.2008,122:1266-1270.
[63]Rouphael, N. G. et al. Clostridium difficile-associated diarrhea:an emerging threat to pregnant women. Am. J. Obstet. Gynecol.2008,198:635. el-e6.
[64]Huang H, Wu S, Wang M, et al. Clostridium difficile infections in a Shanghai hospital: antimicrobial resistance, toxin profiles and ribotype. Int Antimicrob Agents.2009,33(4): 339-342.
[65]Sayedy L, Kothari D, Richards RJ. Toxic megacolon associated Clostridium difficile colitis. World J Gastrointest Endosc.2010,2(8):293-7.
[66]Cheng, V. C., W. C. Yam, J. F. Chan, et al. Clostridium difficile ribotype 027 arrives in HongKong. Int. J. Antimicrob. Agengts.2009,34:492-493.
[67]Alexander Indra, Marion Blaschitz, Silvia Kernbichler, et al. Mechanisms behind variation in the Clostridium difficile 16S-23S rRNA intergenic spacer region. Journal of Medical Microbiology.2010,59:1317-1323.
[68]张维亚,张瑞.克林霉素副作用与艰难梭状芽孢杆菌的关系.中国实用医药.2009,4(30)：131-132.
[69]杨昭徐.PPI引起艰难梭状芽孢杆菌性腹泻的危险因素.中国医院用药评价与分析.2006,3：147-150.
[70]贾晋松,黄晓军,刘代红等.异基因造血干细胞移植过程中患者艰难梭菌感染与肠道微生态失调关系的临床研究.中国实验血液学杂志.2008,16(1)：135-139.
[71]Sandra Janezic, Maja Rupnik. Molecular typing methods for Clostridium difficile: pulsed-field gel electrophoresis and PCR ribotyping. Methods in Molecular Biology.2010, 646:55-65.
[72]Redelings MD, Sorvillo F, Mascola. Increase in Clostridium difficile related mortality rates, United States,1999-2004. Emerging Infectious Diseases.2007,13(9):1417-1419.
[73]J. Freeman, M. P. Bauer, S. D. Baines, et al. The changing epidemiology of clostridium difficile infections. Clinical Microbiology Reviews.2010,23(3):529-549.
[74]Richard A. Stabler, Lisa F. Dawson, Leslie T. H. Phua, et al. Comparative analysis of BI/NAP 1/027 hypervirulent strains reveals novel toxin B-encoding gene (tcdB) sequences. Journal of Medical Microbiology.2008,57:771-775.
[75]Richard A Stabler, Miao He, Lisa Dawson, et al. Comparative genome and phenotypic analysis of dseweez Clostridium difficile 027 strains provides insight into the evolution of a hypervirulent bacterium. Genome Biology.2009,10(9):102.1-102.15.
[76]Spigaglia P, Mastrantonio P. Analysis of macrolide-lincosamide-streptogramin B (MLS(B)) resistance determinant in strains of Clostridium difficile. icrob Drug Resist.2002, 8(1):45-53.
[77]L. Clifford McDonald, George E. Killgore, Angela Thompson, et al. An epidemic, toxin gene-variant strain of Clostridium difficile. The New England Journal of Medicine.2005, 353(23):2433-2441.
[78]Trevor D. Lawley, Simon Clare, Alan W. Walker, et al. Antibiotic treatment of clostridium difficile carrier mice triggers a supershedder state, spore-mediated transmission, and severe disease in immunocompromised hosts. Infection and Immunity.2009,77(9): 3661-3669.
[79]V. K. Viswanathan, M. J. Mallozzi, Gayatri Vedantam. Clostridium diffidicle infection an overview of the disease and its pathogenesis epidemiology and interventions. Gut Microbes. 2010,1(4):234-242.
[80]Yvonne Kleypas, Dennis McCubbin, Hlizabetb S. Curnow. The role of environmental cleaning in health care-associated infections. Critical Care Nursing Quarterly.2011,1(49): 11-17.
[81]Kato H, Kato N, Watanabe K. Relapses or reinfections:analysis of a case of Clostridium difficile-associated colitis by two typing systems. Curr Microbiol.1996,33(4):220-3.
[82]G. L. O'NEILL, M. H. BEAMAN, T. V. RILEY. Relapse versus reinfection with Clostridium diffcile. Epidemiol Infect.1991,107:627-635.
[83]She RC, Durrant RJ, Petti CA. Evaluation of enzyme immunoassays to detect Clostridium difficile toxin from anaerobic stool culture. Am J Clin Pathol.2009,131(1):81-4.
[84]C. Wenisch, B. Parschalk, M. Hasenbundl, A. M. Hirschl, et al. Comparison of vancomycin, teicoplanin, metronidazole, and fusidic acid for the treatment of clostridium difficile-associated diarrhea. Clinical Infectious Diseases.1996,22:813-8.
[85]Sullivan KM, Spooner LM. Fidaxomicin:a macrocyclic antibiotic for the management of Clostridium difficile infection. Ann Pharmacother.2010,44(2):352-9.
[86]Johnson S, Homann SR, Bettin KM, et al. Treatment of asymptomatic Clostridium difficile carriers (fecal excretors) with vancomycin or metronidazole, A randomized, placebo-controlled trial.1992,117(4):297-302.
[87]Alan R. Salkind, Clostridium difficile:an update for the primary care clinician. South Med J.2010,103(9):896-902.
[88]George Tannous, Guy Neff, Nyingi Kemmer. Therapeutic Success of Rifaximin for Clostridium difficile Infection Refractory to Metronidazole and Vancomycin. Gastroenterology.2010,4:404-409.
[89]Shah D, Dang MD, Hasbun R, et al. Clostridium difficile infection:update on emerging antibiotic treatment options and antibiotic resistance. Expert Rev Anti Infect Ther.2010,8(5): 555-64.
[90]Johnson AP. New antibiotics for selective treatment of gastrointestinal infection caused by Clostridium difficile. Expert Opin Ther Pat.2010,20(10):1389-99.
[91]Miller M. Fidaxomicin (OPT-80) for the treatment of Clostridium difficile infection. Expert Opin Pharmacother.2010,11(9):1569-78.
[92]Poxton IR. Fidaxomicin:a new macrocyclic, RNA polymerase-inhibiting antibiotic for the treatment of Clostridium difficile infections. Future Microbiology.2010,5(4):539-548.
[93]Tannock GW, Munro K, Taylor C, et al. A new macrocyclic antibiotic, fidaxomicin (OPT-80), causes less alteration to the bowel microbiota of clostridium difficile-infected patients than does vancomycin. Microbioligy.2010,156(11):3354-9.
[94]Thomas J. Louie, M.D., Mark A. Miller, M.D., Kathleen M. Mullane, et al. Fidaxomicin versus Vancomycin for Clostridium difficile Infection. The New England Journal of Medicine.2011,364:422-431.
[95]John G. Bartlett, Clostridium difficile:progress and challenge, Annals of the New York academy of sciences.2010,1213:62-69.
[96]Lsrael Lowy, Deborah C. Molrine, Brett A. Leav, et al. Treatment with monoclonal antibodies against Clostridium difficile toxins. The New England Journal of Medicine.2010, 362(3):197-205.
[97]Gregory J. Babcock, Teresa J. Broering, Hector J. Hernandez, et al. Human monoclonal antibodies directed against TcdA and TcdB prevent Clostridium difficile-induced mortality in hamsters. Infection and Immunity.2006,74(11):6339-6347.
[98]Tomoaki Saito, Soichiro Kimura, Kazuhiro Tateda, et al. Evidence of intravenous immunoglobulin as a critical supportive therapy against Clostridium difficile toxin-mediated lethality in mice. Journal of Antimicrobial Chemotherapy.2011,66(5):1096-1099.
[99]Hassoun A, Ibrahim F. Use of intravenous immunoglobulin for the treatment of severe Clostridium difficile colitis. Am J Geriatr Pharmacother.2007,5(1):48-51.
[100]Young KW, Munro IC, Taylor SL, et al. The safety of whey protein concentrate derived from the milk of cows immunized against Clostridium difficile, Regul Toxicol Pharmacol. 2007,47(3):317-26.
[101]Numan SC, Veldkamp P, Kuijper EJ. Clostridium difficile-associated diarrhoea:bovine anti-Clostridium difficile whey protein to help aid the prevention of relapses. Gut.2007, 56(6):888-9.
[102]Van Dissel JT, de Groot N, Hensgens CM. Bovine antibody-enriched whey to aid in the prevention of a relapse of Clostridium difficile-associated diarrhoea:preclinical and preliminary clinical data. J Med Microbiol.2005,54(2):197-205.
[103]KarlWeiss, Toxin-binding treatment for Clostridium difficile:a review including reports of studies with tolevamer, International Journal of Antimicrobial Agents.2009,33:4-7.
[104]Sonia S. Yoon, Lawrence J. Brandt. Treatment of Refractory/Recurrent C. difficile-associated Disease by Donated Stool Transplanted Via Colonoscopy A Case Series of 12 Patients. J Clin Gastroenterol.2010,44(8):562-566.
[105]Timothy A. Rubin, Charles E. Gessert, Johannes Aas, et al. Stool transplantation for older patients with Clostridium difficile infection. J Am Geriatr Soc.2009,57(12):2386-7.
[106]Russell G, Kaplan J, Ferraro M, et al. Fecal bacteriotherapy for relapsing Clostridium difficile infection in a child:a proposed treatment protocol. Pediatrics.2010,126(1):239-42.
[107]Von Eichel-Streiber, C. et al. Comparative sequence analysis of the Clostridium difficile toxins A and B, Mol. Gen. Genet.1992,233:260-268.
[108]Kelly CP, LaMont JT. Clostridium difficile---More difficult than ever. N Engl J Med. 2008,359:1932-40.
[109]Stuart H. Cohen, Yajarayma J. Tang, Joseph Silva. Analysis of the Pathogenicity Locus in Clostridium difficile Strains. The Journal of Infectious Diseases.2000,181:659-63.
[110]Maja Rupnik, Mark H. Wilcox, Dale N. Gerding. Clostridium difficile infection:new developments in epidemiology and pathogenesis. Nature Reviews Microbiology.2009, 7:526-536.
[111]I. Just·R. Gerhard. Large clostridial cytotoxins. Rev Physiol Biochem Pharmacol.2004, 152:23-47.
[112]Thomas Jank, Klaus Aktories. Structure and mode of action of clostridial glucosylating toxins:the ABCD model. Trends in Microbiology.2008,16(5):222-229.
[113]Markus Sauerborn, Petra Leukel, Christoph von Eichel-Streiber. The C-terminal ligand-binding domain of Clostridium difficile toxin A (TcdA) abrogates TcdA-specific binding to cells and prevents mouse lethality. FEMS Microbiology Letters.1997,155:45-54.
[114]Hofmann F, Busch C, Prepens U. Localization of the glucosyltransferase activity of Clostridium difficile toxin B to the N-terminal part of the holotoxin. J Biol Chem.1997, 272(17):11074-8.
[115]Thomas Jank, Torsten Giesemann, Klaus Aktories. Rho-glucosylating Clostridium difficile toxins A and B:new insights into structure and function. Clycobiology.2007,17(4): 15-22.
[116]Marinilce F. Santos, Shirley A. McCormack, Zhong Guo. Rho Proteins Play a Critical Role in Cell Migration during the Early Phase of Mucosal Restitution. Journal of Clinical Investigation.1997,100(1):216-255.
[117]Ismael Halabi-Cabezona, Johannes Huelsenbecka, Martin May. Prevention of the cytopathic e□ect induced by Clostridium di□cile Toxin B by active Racl. Federation of European Biochemical Societies.2008,582:3751-3756.
[118]Thomas Jank, Torsten Giesemann, Klaus Aktories. Clostridium difficile Glucosyltransferase Toxin B-essential Amino Acids for Substrate Binding. J Biol Chem.2007, 282 (48):35222-35231.
[119]Esteban Chaves-Olarte, Manfred Weidmann, Christoph von Eichel-Streiber. Toxins A and B from Clostridium difficile Differ with Respect to Enzymatic Potencies, Cellular Substrate Specificities and Surface Binding to Cultured Cells. J. Clin. Invest.1997, 100(7):1734-1741.
[120]Jessica Reineke, Stefan Tenzer, Maja Rupnik. Autocatalytic cleavage of Clostridium difficile toxin B. Nature.2007,446(7134):415-419.
[121]Martina Egerer, Torsten Giesemann, Christian Herrmann. Autocatalytic Processing of Clostridium difficile Toxin B Binding of Inositol Hexakisphosphate. J Biol Chem.2009, 284(6):3389-3395.
[122]Martina Egerer, Torsten Giesemann, Thomas Jank. Auto-catalytic Cleavage of Clostridium difficile Toxins A and B Depends on Cysteine Protease Activity. J Biol Chem. 2007,282(35):25314-25321.
[123]Isa Kreimeyer, Friederike Euler, Alexander Marckscheffel, et al. Autoproteolytic cleavage mediates cytotoxicity of Clostridium difficile toxin A. Naunyn-Schmied Arch Pharmacol.2011,383(3):253-262.
[124]Lehrer RI, Jung G, Ruchala P, et al. Human alpha-defensins inhibit hemolysis mediated by cholesterol-dependent cytolysins. Infect Immun.2009,77(9):4028-40.
[125]Giesemann T, Jank T, Gerhard R, et al. Cholesterol-dependent pore formation of Clostridium difficile toxin A. J Biol Chem.2006,281(16):10808-15.
[126]Selda Genisyuerek, Panagiotis Papatheodorou, Gregor Guttenberg. Structural Determinants for Membrane Insertion, Pore Formation and Translocation of Clostridium difficile Toxin B. Molecular Microbiology.2011,79(6):1643-1654.
[127]Jason G. S. Ho, Antonio Greco, Maja Rupnik, et al. Crystal structure of receptor-binding C-terminal repeats from Clostridium difficile toxin A. PNAS.2005,102 (51):18373-18378.
[128]Cornelia Frisch, Ralf Gerhard, Klaus Aktories. The complete receptor-binding domain of Clostridium difficile toxin A is required for endocytosis. Biochemical and Biophysical Research Communications.2003,300:706-711.
[129]Tanis Dingle, Stefanie Wee, George L Mulvey. Functional properties of the carboxy-terminal host cell-binding domains of the two toxins, TcdA and TcdB, expressed by Clostridium difficile. Glycobiology.2008,18(9):698-706.
[130]Xi Na, Ho Kim,1 Mary P. Moyer, et al. gp96 Is a Human Colonocyte Plasma Membrane Binding Protein for Clostridium difficile Toxin A. Infection and Immunity.2008, 76(7):2862-2871.
[131]M. K. KEEL, J. G. SONGER. The Distribution and Density of Clostridium difficile Toxin Receptors on the Intestinal Mucosa of Neonatal Pigs. Vet Pathol.2007,44:814-822.
[132]Maen Qa'Dan, Matthew Ramsey, Jeremy Daniel, et al. Clostridium difficiletoxin B activates dual caspase-dependent and caspase-independent apoptosis in intoxicated cells. Cellular Microbiology.2002,4(7):425-434.
[133]D. HE, S. J. HAGEN, C. POTHOULAKIS, et al. Clostridium difficile Toxin A Causes Early Damage to Mitochondria in Cultured Cells. gastroenterology.2000,119:139-150.
[134]Ho Kim, Sang Hoon Rhee, Efi Kokkotou, et al. Clostridium difficile Toxin A Regulates Inducible Cyclooxygenase-2 and Prostaglandin E2 Synthesis in Colonocytes via Reactive Oxygen Species and Activation of p38 MAPK. J. Bio.Chem.2005,280(22):21237-21245.
[135]Dan He, Stavros Sougioultzis, Susan Hagen, et al. Clostridium difficile Toxin A Triggers Human Colonocyte IL-8 Release via Mitochondrial Oxygen Radical Generation. gastroenterology.2002,122:1048-1057.
[136]Jin Young Lee, Hyunah Kim, Mi Yeon Cha, et al. Clostridium difficile toxin A promotes dendritic cell maturation and chemokine CXCL2 expression through p38, IKK, and the NF-κB signaling pathway. J Mol Med.2009,87:169-180.
[137]Paola Matarrese, Loredana Falzano, Alessia Fabbri, et al. Clostridium difficile Toxin B Causes Apoptosis in Epithelial Cells by Thrilling Mitochondria Involvement of ATP-Sensitive mitochondrial Potassium Channels. J Biol Chem.2007,282(12):9029-9041.
[138]Pothoulakis C., Lamont J.T., et al. Microbes and microbial toxins:paradigms for microbial-mucosal interactions Ⅱ. The integrated response of the intestine to Clostridium difficile toxins. Am J Physiol Gastrointest Liver Physiol.2001,280:G178-183.
[139]Kelly C.P., Becker S., Linevsky J.K., et al. Neutrophil recruitment in Clostridium difficile toxin A enteritis in the rabbit. J. Clin. Invest.1994,93:1257-1265.
[140]Lyerly D.M., Lockwood D.E., Richardson S.H., et al. Biological activities of toxins A and B of Clostridium difficile. Infect. Immun.1982,35:1147-1150.
[141]Lyerly D.M., Saum K.E., MacDonald D.K., et al. Effects of Clostridium difficile toxins given intragastrically to animals. Infect. Immun.1985,47:349-352.
[142]Mitchell T.J., Ketley J.M., Haslam S.C., et al. Effect of toxin A and B of Clostridium difficile on rabbit ileum and colon. Gut.1986,27:78-85.
[143]Lima A.A., Lyerly D.M., Wilkins T.D.,et al. Effects of Clostridium difficile toxins A and B in rabbit small and large intestine in vivo and on cultured cells in vitro. Infect. Immun.1988, 56:582-588.
[144]Bongaerts, Lyerly. Role of toxins A and B in the pathogenesis of Clostridium difficile disease. Infect. Immun.1994,17:1-12.
[145]Ketley J.M., Mitchell T.J., Candy D.C., et al. The effects of Clostridium difficile crude toxins and toxin A on ileal and colonic loops in immune and non-immune rabbits. J. Med. Microbiol.1987,24:41-52.
[146]Riegler M., Sedivy R., Pothoulakis C., et al. Clostridium difficile toxin B is more potent than toxin A in damaging human colonic epithelium in vitro. J. Clin. Invest.1995,95: 2004-2011.
[147]Savidge T.C., Pan W.H., Newman P, et al. Clostridium difficile toxin B is an inflammatory enterotoxin in human intestine. Gastroenterology.2003,125: 413-420.
[148]Borriello S.P., Wren B.W., Hyde S., et al. Molecular, immunological, and biological characterization of a toxin A-negative, toxin B-positive strain of Clostridium difficile. Infect. Immun.1992,60:4192-4199.
[149]Lyras D., O'Connor J.R., Howarth P.M., et al. Toxin B is essential for virulence of Clostridium difficile. Nature.2009,458:1176-1179.
[150]Geric, B., Carman, Rupnik, et al. Binary toxin-producing, large clostridial toxin-negative Clostridium difficile strains are enterotoxic but do not cause disease in hamsters. J. Infect. Dis.2006,193:1143-1150.
[151]Schwan, C. Stecher, B.Tzivelekidis, et al. Clostridium difficile toxin CDT induces formation of microtubule-based protrusions and increases adherence of bacteria. PLoS Pathog. 2009,5:e1000626.
[152]Qiu B., Pothoulakis C., Castagliuolo I., et al. Participation of reactive oxygen metabolites in Clostridium difficile toxin A-induced enteritis in rats. Am. J. Physiol.1999, 276:G485-G490.
[153]Xingmin Sun, Tor Savidge, Hanping Feng. The Enterotoxcity of Clostridium difficile. Toxins.2010,2:1848-1880.
[154]Qiu B., Pothoulakis C., Castagliuolo I., et al. Nitric oxide inhibits rat intestinal secretion by Clostridium difficile toxin A but not Vibrio cholerae enterotoxin. Gastroenterology.1996, 111:409-418.
[155]Savidge TC, Urvil P, Oezguen N, et al. Host S-nitrosylation inhibits clostridial small molecule-activated glucosylating toxins. Nature Medicine.2011,17(9):1136-41.
[156]Meyer G.K., Neetz A., Brandes, G, et al. Clostridium difficile toxins A and B directly stimulate human mast cells. Infect. Immun.2007,75:3868-3876.
[157]Lima A.A., Nascimento N.R., Fang G.D., et al. Role of phospholipase A2 and tyrosine kinase in Clostridium difficile toxin A-induced disruption of epithelial integrity, histologic inflammatory damage and intestinal secretion.J. Appl. Toxicol.2008,28:849-857.
[158]Hayashi H., Szaszi K., Coady-Osberg N., et al. Inhibition and redistribution of NHE3, the apical Na+/H+exchanger, by Clostridium difficile toxin B. J. Gen. Physiol.2004,123: 491-504.
[159]Alcantare C.S., Jin X.H., Brito G.A., et al. Angiotensin II subtype 1 recptor blockade inhibits Clostridium difficile toxin A-induced intestinal secretion in a rabbit model. J. Infect. Dis.2005,191:2090-2096.
[160]Na X., Zhao D., Koon H.W., et al. Clostridium difficile toxin B activates the EGF receptor and the ERK/MAP kinase pathway in human colonocytes. Gastroenterology.2005, 128:1002-1011.
[161]Calabi E., Fairweather N. Patterns of sequenc conservation in the S-layer proteins and related sequences in Clostridium difficile. J. Bacteriol.002,184:33886-33897.
[162]Drudy D. et al. Human antibody response to surface layer proteins in Clostridium difficile infection. FEMS Immunol. Med. Microbiol.2004,41:237-242.
[163]Wright A. et al. Proteomic analysis of cell surface proteins from Clostridium difficile. Proteomics.2005,5:2443-2452.
[164]Merrigan MM, Gerding DN, Vedantam G. Hypervirulent Clostridium difficile strains have altered protein expression and host-cell adherence. Eighth Biennial Conference of Anaerobe Society of America PI-12 (Boise, Idaho).2006.
[165]Pothoulakis C., Sullivan R., Melnick D.A., et al. Clostridium difficile toxin A stimulates intracellular calcium release and chemotactic response in human granulocytes. Am. J. Pathol. 1988,81:1741-1745.
[166]Ishida Y., Maegawa T., Kondo T., et al. Essential involvement of IFN-gamma in Clostridium difficile toxin A-induced enteritis. J. Immunol.2004,172:3018-3025.
[167]Krivan H.C., Clark G.F., Smith D.F., et al. Cell surface binding site for Clostridium difficile enterotoxin:evidence for a glycoconjugate containing the sequence Gal alpha 1-3Gal beta 1-4GlcNAc. Infect. Immun.1986,53:573-581.
[168]Pothoulakis C., Castagliuolo I., LaMont J.T. Nerves and Intestinal Mast Cells Modulate Responses to Enterotoxins. News Physiol. Sci.1998,13:58-63.
[169]Wershil B.K., Castagliuolo I., Pothoulakis C. Direct evidence of mast cell involvement in Clostridium difficile toxin A-induced enteritis in mice. Gastroenterology.1998,114: 956-964.
[170]Steinman R.M. The dendritic cell system and its role in immunogenicity. Annu. Rev. Immunol.1991,9:271-296.
[171]Lee J.Y., Kim H., Cha M.Y., et al. Clostridium difficile toxin A promotes dendritic cell maturation and chemokine CXCL2 expression through p38, IKK, and the NF-kappaB signaling pathway. J. Mol. Med.2009,87:169-180.
[172]Souza M.H., Melo-Filho A.A., Rocha M.F., et al. The involvement of macrophage-derived tumour necrosis factor and lipoxygenase products on the neutrophil recruitment induced by Clostridium difficile toxin B. Immunology.1997,91:281-288.
[173]Linevsky JK, Pothoulakis C, Keates S, et al. IL-8 release and neutrophil activation by Clostridium difficile toxin-exposed human monocytes. Am J Physiol.1997,273:1333-1340.
[174]Ng J., Hirota S.A., Gross O., et al. Clostridium difficile toxin-induced inflammation and intestinal injury are mediated by the inflammasome. Gastroenterology.2010,139(2):542-552.
[175]Rocha M.F., Maia M.E., Bezerra L.R., et al. Clostridium difficile toxin A induces the release of neutrophil chemotactic factors from rat peritoneal macrophages:role of interleukin-lbeta, tumor necrosis factor alpha, and leukotrienes. Infect. Immun.1997,65: 2740-2746.
[176]Flegel W.A., Muller F., Daubener W., et al. Cytokine response by human monocytes to Clostridium difficile toxin A and toxin B. Infect. Immun.1991,59:3659-3666.
[177]Abrams G.D., Allo M., Rifkin G.D., et al. Mucosal damage mediated by clostridial toxin in experimental clindamycin-associated colitis. Gut.1980,21,493-499.
[178]Czuprynski C.J., Johnson W.J., Balish E., et al. Pseudomembranous colitis in Clostridium difficile-monoassociated rats. Infect. Immun.1983,39:1368-1376.
[179]Fekety R., Silva J., Toshniwal R., et al. Antibiotic-associated colitis:effects of antibiotics on Clostridium difficile and the disease in hamsters. Rev. Infect. Dis.1979,1: 386-397.
[180]Knoop F.C. Clindamycin-associated enterocolitis in guinea pigs:evidence for a bacterial toxin. Infect. Immun.1979,23:31-33.
[181]Chang T.W., Bartlett J.G., Gorbach S.L., et al. Clindamycin-induced enterocolitis in hamsters as a model of pseudomembranous colitis in patients. Infect. Immun.1978,20: 526-529.
[182]Cormier G., Dubos F., Raibaud P. Modulation of cytotoxin production by Clostridium difficile in the intestinal tracts of gnotobiotic mice inoculated with various human intestinal bacteria. Appl. Environ. Microbiol.1985,49:250-252.
[183]Lusk R.H., Fekety R., Silva J., Browne R.A., et al. Clindamycin-induced enterocolitis in hamsters. J. Infect. Dis.1978,137:464-475.
[184]Price A.B., Larson H.E., Crow J. Morphology of experimental antibiotic-associated enterocolitis in the hamster:a model for human pseudomembranous colitis and antibiotic-associated diarrhoea. Gut 1979,20:467-475.
[185]Rehg J.E., Pakes S.P. Implication of Clostridium difficile and Clostridium perfringens iota toxins in experimental lincomycin-associated colitis of rabbits. Lab. Anim. Sci.1982,32: 253-257.
[186]Sugiyama T., Mukai M., Yamashita R., et al. Experimental models of Clostridium difficile enterocolitis in gnotobiotic mice. Prog. Clin. Biol. Res.1985,181:203-206.
[187]Chen X., Katchar K., Goldsmith J.D., et al. A mouse model of Clostridium difficile-associated disease. Gastroenterology 2008,135:1984-1992.
[188]Steele J., Feng H., Parry N., et al. Piglet models of acute or chronic Clostridium difficile illness. J. Infect. Dis.2010,201:428-434.
[189]Hamm E.E., Voth D.E., Ballard J.D. Identification of Clostridium difficile toxin B cardiotoxicity using a zebrafish embryo model of intoxication. Proc. Natl. Acad. Sci.2006, 103:14176-14181.
[190]Rolfe R.D., Iaconis J.P. Intestinal colonization of infant hamsters with Clostridium difficile. Infect. Immun.1983,42:480-486.
[191]M. K. Keel, J. G. Songer. The distribution and density of Clostridium difficile toxin receptors on the intestinal mucosa of neonatal pigs. Vet Pathol.2007,44:814-822.
[192]Hippenstiel S., Schmeck B., N'Guessan P.D., et al. Rho protein inactivation induced apoptosis of cultured human endothelial cells. Am. J. Physiol. Lung Cell. Mol. Physiol.2002, 283, L830-L838.
[193]Sarah A. Kuehnel, Stephen T. Cartmanl, John T. Heap, et al. The role of toxin A and toxin B in Clostridium difficile infection. Nature.2010,467(7316):711-713.
[194]Tim Du MScl, Michelle J Alfa. Translocation of Clostridium difficiletoxin B across polarized Caco-2 cell monolayers is enhanced by toxin A. Can J Infect Dis.2004,15(2): 83-88.
[195]P. A. Sutton, S. Li, J. Webb, et al. Essential role of toxin A in C. difficile 027 and reference strain supernatant-mediated disruption of Caco-2 intestinal epithelial barrier function. Clinical and Experimental Immunology.2008,153:439-447.
[196]Arnaud L, Gorochov G., Charlotte F, Lvovschi V, Parizot C, et al. Systemic perturbation of cytokine and chemokine networks in Erdheim-Chester disease:a single-center series of 37 patients. Blood.2011,117:2783-2790.
[197]Katsikis PD, Mueller YM, Villinger F. The Cytokine Network of Acute HIV Infection: A Promising Target for Vaccines and Therapy to Reduce Viral Set-Point? PLoS Pathog.2011, 7:e1002055.
[198]Cai H, Yu DH, Tian X, Zhu YX. Coadministration of interleukin 2 plasmid DNA with combined DNA vaccines significantly enhances the protective efficacy against Mycobacterium tuberculosis. DNA Cell Biol.2005,24:605-613.
[199]Nambiar JK, Ryan AA, Kong CU, et al. Modulation of pulmonary DC function by vaccine-encoded GM-CSF enhances protective immunity against Mycobacterium tuberculosis infection. Eur J Immunol.2010,40:153-161.
[200]Zhuang Y, Li G, Zhang X, et al. rIL-2 zai xiao shu ti nei kang jie he neng li de yan jiu. Current Immunology.1992,12:3.
[201]Szeliga J, Daniel DS, Yang CH, et al. Granulocyte-macrophage colony stimulating factor-mediated innate responses in tuberculosis. Tuberculosis.2008,88:7-20
[202]Toubaji A, Hill S, Terabe M, et al. The combination of GM-CSF and IL-2 as local adjuvant shows synergy in enhancing peptide vaccines and provides long term tumor protection. Vaccine.2007,25:5882-5891.
[203]Ahlers JD, Dunlop N, Alling DW, et al. Cytokine-in-adjuvant steering of the immune response phenotype to HIV-1 vaccine constructs:granulocyte-macrophage colony-stimulating factor and TNF-alpha synergize with IL-12 to enhance induction of cytotoxic T lymphocytes. J Immunol.1997,158:3947-3958.
[204]Orlova MO, Majorov KB, Lyadova IV, et al. Constitutive differences in gene expression profiles parallel genetic patterns of susceptibility to tuberculosis in mice. Infect Immun.2006, 74:3668-3672.
[205]Ota H, Kumagai S, Morinobu A, et al. Enhanced production of transforming growth factor-beta (TGF-beta) during autologous mixed lymphocyte reaction of systemic sclerosis patients. Clin Exp Immunol.1995,100:99-103.
[206]Dheda K, Chang JS, Breen RA, et al. In vivo and in vitro studies of a novel cytokine, interleukin 4delta2, in pulmonary tuberculosis. Am J Respir Crit Care Med.2005,172: 501-508.
[207]Rial D. Rolfe. Binding kinetics of Clostridium difficile toxins A and B to intestinal brush border membranes from infant and adult hamsters. Infection and Immunity.1991,59(4): 1223-1230.
[208]Barroso LA, Moncrief JS, Lyerly DM, Wilkins TD. Mutagenesis of the Clostridium difficile toxin B and effect on cytotixic activity. Microb Pathog.1994,16(4):297-303.
[209]David Albesa-Jove, Thomas Bertrand, Elisabeth P. Carpenter. Four Distinct Structural Domains in Clostridium difficile Toxin B Visualized Using SAXS. J. Mol. Biol.2010,396: 1260-1270.
[210]Rory N. Pruitt, Melissa G. Chambers, Kenneth K.-S. Ng, et al. Structural organization of the functional domains of Clostridium difficile toxins A and B. PNAS.2010,107(30): 13467-72.
[211]von Eichel-Streiber C, Boquet P, Sauerborn M, et al. Large clostridial cytotoxins-a family of glycosyltransferases modifying small GTP-binding proteins. Trends Microbiol. 1996,4:375-382.
[212]Xingmin Sun, Xiangyun He, Saul Tzipori, et al. Essential role of the glucosyltransferase activity in Clostridium difficile toxin-induced secretion of TNF-a by macrophages. Microbial Pathogenesis.2009,46(6):298-305.
[213]Torsten Giesemann, Gregor Guttenberg, Klaus Aktories, Human a-Defensins Inhibit Clostridium difficile Toxin B. Gastroenterology.2008,134:2049-2058.
[214]Daniel M. Musher, Saima Aslam, Nancy Logan, et al. Relatively poor outcome after treatment of Clostridium difficile colitis with metronidazole. Clin Infect Dis.2005,40(11): 1586-1.
[215]John G. Bartlett. Clostridium difficile:progress and challenges, Annals of the New York Academy of Science.2010,1213:62-69.
[216]Johannes Huelsenbeck, Stefanie Dreger, Ralf Gerhard, et al. Difference in the Cytotoxic Effects of Toxin B from Clostridium difficile Strain VPI 10463 and Toxin B from Variant Clostridium difficile Strain 1470. Inf. Immu.2007,75(2):801-809.
[217]Antonio Greco, Jason G S Ho, Shuang-Jun Lin, et al. Carbohydrate recognition by Clostridium difficile toxin A. Nature Structural & Biology.2006,13(5):460-461.
[218]Chiou-Yueh Yeh, Chun-Nan Lin, Chuan-Fa Chang, et al. C-Terminal Repeats of Clostridium difficile Toxin A Induce Production of Chemokine and Adhesion Molecules in Endothelial Cells and Promote Migration of Leukocytes, Infection and Immunity.2008,76(3): 1170-1178.
[219]Martin Riegler, Roland Sedivy, Charalabos Pothoulakis, Gerhard Hamilton, et al. Clostridium difficile toxin B is more potent than toxin A in damaging human colonic epithelium in vitro. Journal of Clinical Investigation.1995,95:2004-2011.
[220]Jennifer Steele, Kevin Chen, Xingmin Sun, et al. Systemic Dissemination of Clostridium difficile Toxins A and B is Associated with Severe, Fatal Disease in Animal Models, The Journal of Infectious Diseases.2011 (accepted).
© 2004-2018 中国地质图书馆版权所有 京ICP备05064691号 京公网安备11010802017129号 地址：北京市海淀区学院路29号 邮编：100083 电话：办公室：(+86 10)66554848；文献借阅、咨询服务、科技查新：66554700 |