Epsilon Toxin-Producing Clostridium perfringens within the MS Gut Microbiome and Epsilon Toxin Function on Immune Privilege. Epsilon Toxin-Producing Clostridium perfringens within the MS Gut Microbiome and Epsilon Toxin Function on Immune Privilege

Multiple Sclerosis (MS) is a complex disease of the CNS believed to require one or more environmental triggers and is characterized by episodic formation of inflammatory demyelinating lesions in the brain and spinal cord. Gut dysbiosis is a common feature in MS and here, using enhanced and quantitative PCR detection, we show that people with MS are more likely to harbor and have higher abundance of epsilon toxin (ETX)-producing strains of Clostridium perfringens within their gut microbiome compared to healthy controls (HC). MS patient-derived isolates produce functional ETX and have a genetic architecture typical of highly conjugative plasmids. In the active immunization model of experimental autoimmune encephalomyelitis (EAE), where pertussis toxin (PTX) is used to overcome CNS immune privilege, we find that ETX can substitute for PTX in disease induction. In contrast to PTX-induced EAE, where inflammatory demyelination is largely restricted to the spinal cord, ETX-induced EAE results in multifocal demyelination in the corpus callosum, thalamus, cerebellum, brainstem, and spinal cord, more akin to the lesion distribution observed in MS. Transcriptional profiles from CNS endothelial cells reveal ETX-induced genes that are known to play a role in overcoming CNS immune privilege. Together, these findings support ETX-producing strains of C. perfringens as biologically plausible pathogens in MS to trigger inflammatory demyelination in the context of circulating myelin autoreactive lymphocytes. Overall design: Comparative gene expression profiling analysis of RNA-seq data of CNS endothelial cells treated with PBS, ETX (0.5 μg/kg b.w.), or PTX (5 μg/kg b.w).

多发性硬化症（Multiple Sclerosis, MS）是一种复杂的中枢神经系统（Central Nervous System, CNS）疾病，被认为需要一种或多种环境触发因素，其特征为脑与脊髓内反复出现炎症性脱髓鞘病灶。肠道菌群失调是MS患者的常见特征。本研究通过增强型定量聚合酶链反应（quantitative PCR, PCR）检测发现，与健康对照（Healthy Controls, HC）相比，MS患者肠道微生物组中更易定植产ε毒素（epsilon toxin, ETX）的产气荚膜梭菌（Clostridium perfringens）菌株，且其丰度更高。从MS患者体内分离得到的菌株可产生活性ETX，且其遗传结构具备高接合性质粒的典型特征。在采用百日咳毒素（pertussis toxin, PTX）突破中枢神经系统免疫豁免的实验性自身免疫性脑脊髓炎（Experimental Autoimmune Encephalomyelitis, EAE）主动免疫模型中，本研究发现ETX可在疾病诱导过程中替代PTX发挥作用。与PTX诱导的EAE（其炎症性脱髓鞘主要局限于脊髓）不同，ETX诱导的EAE可在胼胝体、丘脑、小脑、脑干及脊髓内引发多灶性脱髓鞘，其病灶分布与MS患者的病灶分布更为相似。对中枢神经系统内皮细胞的转录组谱分析显示，ETX可诱导表达一系列已知可突破中枢神经系统免疫豁免的基因。综合上述研究结果，本研究支持产ETX的产气荚膜梭菌菌株可作为MS的生物学合理病原体，在循环髓鞘反应性淋巴细胞存在的情况下触发炎症性脱髓鞘病变。实验设计概述：对经磷酸盐缓冲液（Phosphate Buffered Saline, PBS）、ETX（0.5 μg/kg 体重）或PTX（5 μg/kg 体重）处理的中枢神经系统内皮细胞的RNA测序（RNA-sequencing, RNA-seq）数据进行比较基因表达谱分析。