CN114085798A - 一种植物乳杆菌及其在预防和/或治疗疾病中的应用 - Google Patents
一种植物乳杆菌及其在预防和/或治疗疾病中的应用 Download PDFInfo
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- CN114085798A CN114085798A CN202111622634.7A CN202111622634A CN114085798A CN 114085798 A CN114085798 A CN 114085798A CN 202111622634 A CN202111622634 A CN 202111622634A CN 114085798 A CN114085798 A CN 114085798A
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Abstract
本发明公开了一种植物乳杆菌及其在预防和/或治疗疾病中的应用,所述的疾病为炎症性肠病和/或中枢神经***疾病。本发明分离得到植物乳杆菌(Lactobacillus plantarum)Pm002,保藏于中国典型培养物保藏中心(CCTCC)、保藏编号为CCTCC NO:M 2021927。经由活体内及体外的相关实验证实,植物乳杆菌Pm002可以抑制肠道有害菌生长,改善肠道菌群,对炎症性肠病具有优异的抵抗力;并且还发现Pm002对于中枢神经***炎症性脱髓鞘疾病具有明显的治疗和改善作用,这预示植物乳杆菌Pm002菌具有治疗炎症性肠病和多发性硬化(MS)的前景,具有重要的应用价值。
Description
技术领域
本发明涉及微生物技术领域,具体涉及一种植物乳杆菌Pm002及其在预防和/或治疗疾病中的应用,所述的疾病为炎症性肠病和/或中枢神经***疾病。
背景技术
炎症性肠病(Inflammatory bowel disease,IBD)为累及回肠、直肠、结肠的一种特发性肠道炎症性疾病。临床表现腹泻、腹痛,甚至可有血便。溃疡性结肠炎(ulcerativecolitis,UC)与克罗恩病(CD)是IBD的两种主要类型。目前IBD的病因及发病机制尚不明确,多认为是环境因素作用于易感人群引发肠免疫紊乱,最终导致肠道炎症反应形成。来自流行病学、临床及实验动物模型的研究显示IBD患者存在肠道菌群的异常,而肠道菌群是这种免疫损伤过程的重要触发因子。目前临床上通过改变肠道菌群的方法主要包括使用抗生素、益生菌以及进行粪菌移植,抗生素治疗有利于CD患者结肠炎症反应的缓解、脓肿并发症的治疗及预防术后复发,但对UC患者无效,且长期应用存在较大不良反应。粪菌移植(fecalmicrobiota transplantation,FMT)是复发性难辨梭状芽胞杆菌感染的有效治疗手段,但对于FMT在IBD治疗的有效性,研究结果并不一致。
多发性硬化症(multiple sclerosis,MS)是最常见的中枢神经***(centralnervoussystem,CNS)自身免疫性疾病,属慢性炎症性脱髓鞘性疾病,病理特征为白质有T淋巴细胞、巨噬细胞的浸润和局部脱髓鞘病变,临床表现为行动能力受损、视力障碍、疲乏和认知损害。多发性硬化应及早诊断,及早治疗,遵循循证医学证据,结合患者的经济条件和意愿,进行合理治疗。但是目前有循证医学论据支持的药物相当有限,并且价格昂贵。目前国内治疗MS因受经济条件和医疗保险报销等因素限制,只有极少数患者能用干扰素β治疗,绝大多数还是使用肾上腺糖皮质激素、免疫抑制剂等治疗,效果有限并且副作用大。
本发明筛选得到一株植物乳杆菌Pm002,并且通过实验发现植物乳杆菌Pm002能够显著抑制肠道有害菌生长,改善肠道菌群,对炎症性肠病(包括溃疡性结肠炎或克罗恩病)具有优异的抵抗力;而且还发现植物乳杆菌Pm002对于中枢神经***炎症性脱髓鞘疾病(包括多发性硬化、视神经脊髓炎或急性散播性脑脊髓炎)具有明显的治疗和改善作用,这预示植物乳杆菌Pm002菌具有治疗炎症性肠病和多发性硬化(MS)的前景,具有重要的应用价值。
发明内容
为克服现有技术的不足,本发明提供一种植物乳杆菌及其在预防和/或治疗疾病中的应用,所述的疾病为炎症性肠病和/或中枢神经***疾病。
在本发明第一方面,提供一种植物乳杆菌Pm002,其保藏号为CCTCC NO:M2021927。
在本发明第二方面,提供一种组合物,其包含植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物,以及一种或多种辅料。
特别是,该组合物包含植物乳杆菌Pm002以及一种或多种辅料。
具体地,在该组合物中,植物乳杆菌Pm002为活菌或非活菌,特别是活菌。
具体地,该组合物还可以含有其他益生菌和/或益生元,例如,其他对调节肠道菌群稳态,预防和治疗炎症性肠病等肠道疾病的发生有益的微生物和/或益生元。
在本发明的一个实施方案中,该组合物为药物组合物,其包含植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物(特别是植物乳杆菌Pm002活菌),以及一种或多种药学上可接受的辅料。
具体地,该组合物可以选自:药物组合物、食品组合物、保健品组合物、饲料添加剂组合物、饲料组合物等。
具体地,上述药物组合物包含植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物(特别是植物乳杆菌Pm002活菌),以及一种或多种药学上可接受的辅料。
具体地,该药学上可接受的辅料可以为,例如,但不限于,填充剂、粘合剂、润湿剂、崩解剂、润滑剂、矫味剂、香味剂、着色剂、包衣剂、酸度调节剂、防腐剂、稀释剂、悬浮液稳定剂等中的一种或多种。
具体地,该药物组合物可以为任何合适的剂型,特别是口服剂型,例如,液体制剂(如乳剂、混悬剂、糖浆剂等)、固体制剂(如粉剂、片剂、丸剂、颗粒剂、胶囊剂、锭剂等),等。
在本发明的一些实施例中,该药物组合物为液体制剂,其中植物乳杆菌Pm002为活菌,其有效活菌数≥1×109CFU/mL,例如,1×109至10×109CFU/mL,5×109CFU/mL。
具体地,上述食品组合物为人类日常摄取的食品,其可以为直接添加植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物(特别是植物乳杆菌Pm002活菌)的食品组合物,也可以为添加植物乳杆菌Pm002后经过发酵生产的食品组合物。
具体地,该食品组合物可以为乳制品(如发酵乳、酸奶、奶油、干酪等)、豆制品(如豆乳、豆腐乳、豆豉、豆酱等)、果蔬制品(如果蔬饮料、腌制产品(如泡菜、酸菜))、肉制品、调味品等。
具体地,该食品组合物的形式可以采用任意的形式,如,糖果(如压片糖果、凝胶糖果、胶基糖果等)、固体饮料(如粉末、颗粒等)、液体饮料等。
具体地,上述保健品组合物,其能调节人体的机能,但不以治疗疾病为目的,其包含植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物(特别是植物乳杆菌Pm002活菌),以及一种或多种保健品辅料。
具体地,该保健品辅料可以为,例如,但不限于,填充剂、粘合剂、润湿剂、崩解剂、润滑剂、矫味剂、香味剂、着色剂、包衣剂、酸度调节剂、防腐剂、稀释剂、悬浮液稳定剂等中的一种或多种。
具体地,该保健品组合物的形式可以采用任意的形式,如,片剂、丸剂、胶囊剂、糖果(如压片糖果、凝胶糖果、胶基糖果等)、固体饮料(如粉剂、颗粒剂等)、液体饮料等。
在本发明另一个实施方案中,该组合物为饲料添加剂组合物,其为在饲料(如家畜饲料、家畜饮用水、宠物日粮、宠物零食、宠物饮用水等)生产加工、使用过程中以少量或微量添加的物质,其包含植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物(特别是植物乳杆菌Pm002活菌),以及一种或多种添加剂辅料。
具体地,该添加剂辅料可以为,例如,但不限于,填充剂、粘合剂、润湿剂、崩解剂、润滑剂、矫味剂、香味剂、着色剂、包衣剂、酸度调节剂、防腐剂、稀释剂、悬浮液稳定剂等中的一种或多种。
具体地,该饲料添加剂组合物的形式可以采用任意的形式,如,片剂、丸剂、胶囊剂、粉剂、颗粒剂、液体剂等。
具体地,上述饲料组合物为人类日常喂饲动物(例如宠物、家畜、实验动物)的饲料,例如,家畜饲料、宠物日粮、宠物零食等,其可以为直接添加植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物(特别是植物乳杆菌Pm002活菌)的饲料组合物,也可以为添加植物乳杆菌Pm002后经过发酵生产的饲料组合物。
在本发明第三方面,提供植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物在制备预防和/或治疗肠道疾病的药物中的应用。具体地,该肠道疾病可以选自十二指肠、空肠、回肠、盲肠、结肠或直肠疾病,特别是累及回肠、直肠、结肠的肠道疾病。
在本发明的一个实施方案中,该肠道疾病为炎症性肠病,例如,但不限于,溃疡性结肠炎(UC)、克罗恩病(CD)。
具体地,该炎症性肠病可以为慢性炎症性肠病或急性炎症性肠病,特别是急性炎症性肠病。
在本发明另一个实施方案中,该肠道疾病为肠道肿瘤,例如,但不限于,结肠癌、直肠癌。
具体地,该肠道疾病的症状选自:腹痛、腹泻、便血、呕吐、里急后重、隐窝脓肿,以及体重减轻、发热、贫血、低蛋白血症、电解质异常、息肉、肠穿孔等中的一种或多种。
具体地,治疗作用可以为,改善上述肠道疾病症状中的一种或多种,降低肠道疾病的复发率,甚至完全治愈该肠道疾病。
具体地,预防作用可以为,降低肠道疾病的发生率,甚至完全避免患该肠道疾病。
在本发明第四方面,提供植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物在制备预防和/或治疗中枢神经***疾病的药物中的应用。
具体地,该中枢神经***疾病为中枢神经***炎症性脱髓鞘疾病。
具体地,所述中枢神经***炎症性脱髓鞘疾病为多发性硬化、视神经脊髓炎或急性散播性脑脊髓炎,优选为多发性硬化。
具体地,所述多发性硬化包括复发缓解型MS、继发进展型MS、原发进展型MS和进展复发型MS。
具体地,该多发性硬化的症状选自:疲劳、痉挛、共济失调、虚弱、膀胱和肠紊乱、性功能障碍、疼痛、颤动、阵发性表现、视觉障碍、心理问题和认知障碍中的一种或多种。
具体地,所述植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物的治疗作用可以为,改善上述中枢神经***疾病症状中的一种或多种,降低中枢神经***疾病的复发率,甚至完全治愈中枢神经***疾病。
具体地,所述植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物的预防作用可以为,改善上述中枢神经***疾病症状中的一种或多种,降低中枢神经***疾病的发生率,甚至完全避免患中枢神经***疾病。
在本发明第五方面,提供一种预防和/或治疗肠道疾病的方法,其包括向有此需求的受试者施用植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物(特别是植物乳杆菌Pm002活菌),或本发明第二方面所述药物组合物的步骤。
具体地,该肠道疾病可以选自十二指肠、空肠、回肠、盲肠、结肠或直肠疾病,特别是累及回肠、直肠、结肠的肠道疾病。
在本发明的一个实施方案中,该肠道疾病为炎症性肠病,例如,但不限于,溃疡性结肠炎(UC)、克罗恩病(CD)。
在本发明另一个实施方案中,该肠道疾病为肠道肿瘤,例如,但不限于,结肠癌、直肠癌。
具体地,该肠道疾病的症状选自:腹痛、腹泻、便血、呕吐、里急后重、隐窝脓肿,以及体重减轻、发热、贫血、低蛋白血症、电解质异常、息肉、肠穿孔等中的一种或多种。
具体地,治疗作用可以为,改善上述肠道疾病症状中的一种或多种,降低肠道疾病的复发率,甚至完全治愈该肠道疾病。
具体地,预防作用可以为,降低肠道疾病的发生率,甚至完全避免患该肠道疾病。
具体地,该受试者可以为哺乳动物,特别是人类。
在本发明第六方面,提供一种预防和/或治疗中枢神经***疾病的方法,其包括向有此需求的受试者施用植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物(特别是植物乳杆菌Pm002活菌)或包含植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物(特别是植物乳杆菌Pm002活菌)的药物。
具体地,所述中枢神经***疾病为中枢神经***炎症性脱髓鞘疾病。
具体地,所述中枢神经***炎症性脱髓鞘疾病为多发性硬化、视神经脊髓炎或急性散播性脑脊髓炎,优选为多发性硬化。
具体地,所述多发性硬化包括复发缓解型MS、继发进展型MS、原发进展型MS和进展复发型MS。
具体地,该多发性硬化的症状选自:疲劳、痉挛、共济失调、虚弱、膀胱和肠紊乱、性功能障碍、疼痛、颤动、阵发性表现、视觉障碍、心理问题和认知障碍中的一种或多种。
具体地,该受试者可以为哺乳动物,特别是人类。
在本发明第七方面,提供一种调节肠道菌群稳态的方法,其包括向有此需求的受试者施用植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物(特别是植物乳杆菌Pm002活菌),或本发明第二方面所述食品组合物、保健品组合物或饲料添加剂组合物、饲料组合物的步骤。
具体地,该调节肠道菌群稳态的方法为非疾病预防或治疗目的。
具体地,该受试者可以为哺乳动物,例如,人类、家畜(如猪、牛、羊、马、驴、狐、貉、貂、骆驼等)、宠物(如狗、猫、兔、鼠(如豚鼠、仓鼠、沙鼠、龙猫、松鼠等))、实验动物(如猴、狗、兔、猫、鼠(如小鼠、大鼠)等)等,特别是人类。
本发明分离得到植物乳杆菌Pm002,其可以抑制肠道有害菌生长,改善肠道菌群,对炎症性肠病,特别是,溃疡性结肠炎或克罗恩病,具有优异的抵抗力,且无毒副作用;而且,研究还显示,植物乳杆菌Pm002对中枢神经***炎症性脱髓鞘疾病,尤其是多发性硬化、视神经脊髓炎或急性散播性脑脊髓炎同样具有优异的抵抗力,对于实验性自身免疫性脑脊髓炎(EAE)具有明显的治疗和改善作用,具有重要的应用价值。本发明为炎症性肠病和多发性硬化的治疗提供新的药物选择和思路。
本发明中所述植物乳杆菌(Lactobacillus plantarum)Pm002已于2021年7月23日保藏于中国典型培养物保藏中心(地址:中国武汉,武汉大学),保藏编号:CCTCC NO:M2021927。
附图说明
图1所示为生理盐水-DSS对照组、植物乳杆菌Pm002-DSS处理组和鼠李糖乳杆菌LGG-DSS对照组小鼠在DSS诱导肠炎期间体重变化曲线。
图2所示为生理盐水-DSS对照组、植物乳杆菌Pm002-DSS处理组和鼠李糖乳杆菌LGG-DSS对照组小鼠在DSS诱导肠炎期间粪便评分曲线。
图3所示为生理盐水-DSS对照组、植物乳杆菌Pm002-DSS处理组和鼠李糖乳杆菌LGG-DSS对照组小鼠第11天小鼠结肠的长度。
图4所示为生理盐水-DSS对照组、植物乳杆菌Pm002-DSS处理组和鼠李糖乳杆菌LGG-DSS对照组小鼠远端结肠H&E染色显示肠道上皮破坏情况。
图5为生理盐水-EAE对照组和植物乳杆菌Pm002-EAE处理组小鼠在EAE发病过程中的临床评分。
图6为生理盐水-DSS处理组和植物乳杆菌Pm002-EAE处理组小鼠脊髓组织H&E和Luxol fast blue免疫组织化学染色结果。
图7为生理盐水-DSS处理组和植物乳杆菌Pm002-EAE处理组小鼠脊髓中炎症相关基因mRNA的表达水平结果。
具体实施方式
除非另有定义,本发明中所使用的所有科学和技术术语具有与本发明涉及技术领域的技术人员通常理解的相同的含义。
如本发明中所用,术语“培养物”是指一定时间一定空间内微生物的细胞群或生长物,特别是指经人工接种和培养后,生长有微生物(植物乳杆菌Pm002)群体的液体或固体培养基。
如本发明中所用,术语“裂解物”是指微生物(植物乳杆菌Pm002)细胞裂解后所得产物。
如本发明中所用,术语“菌悬液”是指含有微生物(植物乳杆菌Pm002)细胞的悬浮液。
如本发明中所用,术语“治疗”包括在疾病发作之后,根除、移除、逆转、缓解、改变或控制疾病。
如本发明中所用,术语“预防”指在疾病发作之前,通过治疗以避免、最小化或令疾病或状况难于发作或发展的能力。
如本发明中所用,术语“患者”或“受试者”等等在本文中可交换使用,是指根据本文所述的方法处理的任何动物或其细胞,不论是体外或原位。具体地,前述动物包括哺乳动物,例如,人类、家养动物(即由人类饲养驯化,且可以人为控制其繁殖的动物,用于例如食用、劳役、毛皮、宠物、实验等功能,例如经济动物、宠物、实验动物等),特别是人类。
本文所引用的各种出版物、专利和公开的专利说明书,其公开内容通过引用整体并入本文。
下面将结合本发明实施例,对本发明的技术方案进行清楚、完整地描述,显然,所描述的实施例仅是本发明一部分实施例,而不是全部的实施例。基于本发明中的实施例,本领域普通技术人员在没有作出创造性劳动前提下所获得的所有其他实施例,都属于本发明保护的范围。
实施例1:植物乳杆菌Pm002可以减轻小鼠DSS诱导肠炎表型
(1)准备WT小鼠
从上海灵畅公司购买30只8周龄的野生型BL/C57小鼠,饲养于SPF屏障***内,适应一周后将小鼠随机分成3组,分别为生理盐水-DSS对照组(阴性对照组)、植物乳杆菌Pm002-DSS处理组和鼠李糖乳杆菌LGG-DSS对照组(阳性对照组)。
(2)植物乳杆菌Pm002肠道定植
将植物乳杆菌Pm002和鼠李糖乳杆菌LGG接种于液态MRS培养基中,于37℃条件下厌氧、静置培养20-22h,然后8000rpm离心5min收集菌体,用生理盐水洗涤两次后,再将细菌重悬于生理盐水中,终浓度约5×109CFU/mL。按照每只小鼠200μL菌的剂量灌胃处理WT小鼠,隔天重复处理,持续2周,同时生理盐水-DSS组仅进行生理盐水灌胃处理。2周的灌胃处理结束后,小鼠即可进行诱导肠炎实验,诱导肠炎期间仍隔天灌胃相同菌量。
(3)建立小鼠DSS诱导肠炎模型
将生理盐水-DSS对照组、植物乳杆菌Pm002-DSS处理组和鼠李糖乳杆菌LGG-DSS对照组小鼠饮水换成3%(质量%)葡聚糖硫酸钠(DSS)水溶液,处理9天后换成正常饮水至实验结束。即,DSS诱导肠炎的第1-9天,生理盐水-DSS对照组、植物乳杆菌Pm002-DSS处理组和鼠李糖乳杆菌LGG-DSS对照组小鼠饮用3%DSS水溶液,第10-12天饮用普通饮用水。
诱导肠炎的第1-12天记录体重变化、粪便干稀度、便血情况。粪便评分:0,固态大便;1,固态大便,易变形;2,不成形大便;3,液体状大便。体重变化的对比如图1所示。粪便评分的对比如图2所示。
根据该图可得知,生理盐水-DSS对照组、植物乳杆菌Pm002-DSS处理组和鼠李糖乳杆菌LGG-DSS对照组小鼠饮用DSS后均出现体重减轻,说明DSS肠炎诱导是成功的。进一步比较,植物乳杆菌Pm002-DSS处理组和鼠李糖乳杆菌LGG-DSS对照组小鼠体重减轻幅度显著小于生理盐水-DSS对照组,植物乳杆菌Pm002-DSS处理组和鼠李糖乳杆菌LGG-DSS对照组小鼠体重减轻幅度差异不显著;同时植物乳杆菌Pm002-DSS处理组和鼠李糖乳杆菌LGG-DSS对照组小鼠粪便评分显著低于生理盐水-DSS组,植物乳杆菌Pm002-DSS处理组和鼠李糖乳杆菌LGG-DSS对照组小鼠粪便评分差异不显著,说明植物乳杆菌Pm002-DSS处理组和鼠李糖乳杆菌LGG-DSS对照组小鼠肠炎症状较生理盐水-DSS组有显著改善。
DSS处理第12天,颈椎脱臼法处死小鼠,取出小鼠结肠部分,测量其长度,所得结果如图3所示。取远端结肠组织1cm左右肠段进行***固定,并后续进行HE染色和组织形态分析,所得结果如图4所示。
根据图3,可得知:DSS诱导肠炎模型中,植物乳杆菌Pm002-DSS处理组和鼠李糖乳杆菌LGG-DSS对照组小鼠肠道长度分别为(6.44±0.15cm)和(6.52±0.53cm),显著长于生理盐水-DSS组小鼠(5.2±0.35cm);根据图4,可得知:植物乳杆菌Pm002-DSS组和鼠李糖乳杆菌LGG-DSS对照组小鼠肠道上皮细胞仍可以看到较完整的隐窝结构,而生理盐水-DSS组小鼠肠道上皮则完全被破坏,无完整的隐窝结构。此图从形态学上反映出植物乳杆菌Pm002-DSS组小鼠肠炎发病显著降低。
实施例2:植物乳杆菌Pm002菌可以减轻小鼠EAE模型的表型(1)准备WT小鼠
从上海灵畅公司购买20只8周龄的野生型BL/C57小鼠,饲养于SPF屏障***内,适应一周后将小鼠随机分成2组,分别为生理盐水-EAE对照组和植物乳杆菌Pm002菌-EAE组。
(2)植物乳杆菌Pm002菌肠道定植
将植物乳杆菌Pm002菌接种于液态MRS培养基中,于37℃条件下厌氧、静置培养20-22h,然后8000rpm离心5min收集菌体,用生理盐水洗涤两次后,再将细菌重悬于生理盐水中,终浓度约5×109CFU/mL。按照每只小鼠200μL植物乳杆菌Pm002的剂量灌胃处理WT小鼠,隔天重复处理,持续2周,同时生理盐水-DSS组仅进行生理盐水灌胃处理。
2周的灌胃处理结束后,小鼠即可进行诱导EAE实验。
(3)建立小鼠EAE模型
在弗氏不完全佐剂(IFA)中加入结核分枝杆菌(TB)配成终浓度为5mg/mL的完全弗氏佐剂(complete Freundadjuvant,CFA),髓鞘少突胶质细胞糖蛋白35-55(MOG35-55)肽段用PBS配成5mg/mL。CFA和MOG35-55溶液按水和油体积比1∶1混合,用注射器反复抽推制成油包水样乳白色混悬液。抗原乳剂按0.4mL/只分4点脊柱两侧皮下注射,免疫当天记录为免疫后第0天。各组小鼠于免疫当天和48h分别尾静脉注射200ng/只PTX作为免疫增强剂。
EAE的临床评分标准为:0分,无症状;1分,尾巴无力;2分,尾麻痹及轻微后肢无力;3分,尾麻痹及重度后肢无力;4分,尾麻痹及四肢麻痹;5分,濒死或死亡。临床评分对比如图5所示。
(4)染色分析
小鼠麻醉后1×PBS灌注,解剖取出小鼠的脑与脊髓腰部膨大处,4%多聚甲醛固定,30%蔗糖脱水,冰冻切片,NG2抗体免疫组织化学染色;主动免疫MOG35-55或过继转移Th1/Th17后第20天,EAE小鼠麻醉后1×PBS灌注,解剖取出小鼠脊髓腰部膨大处,4%多聚甲醛固定,石蜡包埋做病理切片,进行HE染色、勒克司坚牢蓝(Luxolfast blue,LFB)染色。组织化学染色分析对比如图6所示。
(5)qPCR检测
EAE小鼠麻醉后1×PBS灌注,解剖取出小鼠脊髓,TRIzol裂解,提取总RNA,再用PrimeScrip RT reagent Kit反转录成cDNA,SYBR Green Master Mix进行real-time PCR(测定基因引物见表1)。炎症因子基因qPCR分析对比如图7所示。
表1 qPCR引物序列
本发明建立了多发性硬化的动物模型EAE小鼠,通过分析小鼠EAE发病过程中的临床评分、组织化学和炎症相关基因mRNA的表达水平等指标,证明植物乳杆菌Pm002能有效缓解动物模型EAE小鼠的发病程度及降低发病率,这是首次发现植物乳杆菌有效缓解动物模型EAE小鼠的发病程度的功能。
根据图5-7显示,生理盐水-EAE对照组小鼠高峰临床得分显著高于植物乳杆菌Pm002菌-EAE组小鼠;H&E染色结果发现生理盐水-EAE对照组小鼠和植物乳杆菌Pm002菌-EAE组小鼠均有不同程度的炎症细胞浸润,但对照组小鼠较严重;LFB染色结果显示生理盐水-EAE对照组小鼠较植物乳杆菌Pm002菌-EAE组小鼠脱髓鞘严重。以上结果说明植物乳杆菌Pm002可以减轻小鼠EAE模型的表型,即改善EAE模型小鼠尾巴或四肢出现的无力或麻痹症状,减轻炎症细胞对小鼠的侵害,缓解EAE模型小鼠的脱髓鞘疾病症状,总的来说,植物乳杆菌Pm002能够有效缓解多发性硬化动物模型EAE小鼠的发病程度以及显著降低动物模型EAE小鼠的发病率,进而使小鼠的脱髓鞘疾病症状得到有效改善。
以上所述仅为本发明的较佳实施例而已,并不用以限制本发明,凡在本发明的精神和原则之内,所作的任何修改、等同替换等,均应包含在本发明的保护范围之内。
本发明中描述的前述实施例和方法可以基于本领域技术人员的能力、经验和偏好而有所不同。
本发明中仅按一定顺序列出方法的步骤并不构成对方法步骤顺序的任何限制。
Claims (13)
1.一种植物乳杆菌Pm002,其保藏编号为CCTCC NO:M 2021927。
2.一种药物组合物,其包含权利要求1所述的植物乳杆菌Pm002,以及一种或多种药学上可接受的辅料。
3.如权利要求2所述的药物组合物,其特征在于,所述药学上可接受的辅料选自:填充剂、粘合剂、润湿剂、崩解剂、润滑剂、矫味剂、香味剂、着色剂、包衣剂、酸度调节剂、防腐剂、稀释剂、悬浮液稳定剂中的一种或多种;
优选地,所述药物组合物为口服剂型,选自:乳剂、混悬剂、糖浆剂、粉剂、片剂、丸剂、颗粒剂、胶囊剂和锭剂;
更优选地,所述药物组合物为液体制剂,其中植物乳杆菌Pm002为活菌,其有效活菌数≥1×109CFU/mL。
4.一种食品组合物,其包含权利要求1所述的植物乳杆菌Pm002。
5.如权利要求4所述的食品组合物,其特征在于,所述食品组合物为直接添加植物乳杆菌Pm002的食品组合物,或添加植物乳杆菌Pm002后经过发酵生产的食品组合物;
优选地,所述食品组合物为乳制品、豆制品、果蔬制品、肉制品或调味品;
优选地,所述食品组合物为糖果、固体饮料或液体饮料。
6.一种保健品组合物,其包含权利要求1所述的植物乳杆菌Pm002,以及一种或多种保健品辅料。
7.如权利要求6的保健品组合物,其特征在于,所述保健品辅料选自:填充剂、粘合剂、润湿剂、崩解剂、润滑剂、矫味剂、香味剂、着色剂、包衣剂、酸度调节剂、防腐剂、稀释剂、悬浮液稳定剂中的一种或多种;
优选地,所述保健品组合物为片剂、丸剂、胶囊剂、糖果、固体饮料、液体饮料。
8.一种饲料添加剂组合物,其包含权利要求1所述的植物乳杆菌Pm002,以及一种或多种添加剂辅料。
9.权利要求1所述的植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物在制备预防和/或治疗肠道疾病的药物中的应用;
优选地,所述肠道疾病为炎症性肠病或肠道肿瘤;
更优选地,所述炎症性肠病为溃疡性结肠炎或克罗恩病。
10.一种调节肠道菌群稳态的方法,其包括向有此需求的受试者施用权利要求1所述的植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物,或权利要求4或5所述的食品组合物,或权利要求6或7所述的保健品组合物,或权利要求8所述的饲料添加剂组合物的步骤。
11.权利要求1所述的植物乳杆菌Pm002、其培养物、其菌悬液或其裂解物在制备预防和/或治疗中枢神经***疾病的药物中的应用。
12.如权利要求11所述的应用,其特征在于,所述中枢神经***疾病为中枢神经***炎症性脱髓鞘疾病。
13.如权利要求12所述的应用,其特征在于,所述中枢神经***炎症性脱髓鞘疾病为多发性硬化、视神经脊髓炎或急性散播性脑脊髓炎,优选为多发性硬化。
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CN101325960A (zh) * | 2005-10-06 | 2008-12-17 | 普罗比公司 | 乳酸杆菌用于治疗自身免疫性疾病的用途 |
US20090208469A1 (en) * | 2005-10-06 | 2009-08-20 | Probi Ab | Use of Lactobacillus for Treatment of Autoimmune Diseases |
CN105106246A (zh) * | 2015-08-20 | 2015-12-02 | 江南大学 | 一种植物乳杆菌zs2058及其用途 |
CN107619811A (zh) * | 2017-11-08 | 2018-01-23 | 光明乳业股份有限公司 | 植物乳杆菌ccfm200菌株与应用 |
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US20090208469A1 (en) * | 2005-10-06 | 2009-08-20 | Probi Ab | Use of Lactobacillus for Treatment of Autoimmune Diseases |
CN105106246A (zh) * | 2015-08-20 | 2015-12-02 | 江南大学 | 一种植物乳杆菌zs2058及其用途 |
CN107619811A (zh) * | 2017-11-08 | 2018-01-23 | 光明乳业股份有限公司 | 植物乳杆菌ccfm200菌株与应用 |
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