肠-肝轴与非酒精性脂肪性肝病

丁佳，吴健

复旦大学基础医学院医学分子病毒学教育部/卫生部重点实验室，上海 200032

随着肥胖、2型糖尿病和代谢综合征的发病率逐年升高，非酒精性脂肪性肝病(non-alcoholic fatty liver disease，NAFLD)已成为20世纪最常见的慢性肝脏疾病之一。NAFLD包括单纯性肝脂肪变、非酒精性脂肪性肝炎(non-alcoholic steatohepatitis，NASH)伴或不伴肝纤维化，后者可进展为终末期肝病(肝硬化)，并在此基础上发生肝细胞肝癌(简称肝癌)。NASH已成为近年来西方国家肝癌发病率升高的主要因素之一。除遗传基因、饮食环境和生活方式等因素与NAFLD的发病相关外，近期研究表明肠道菌群组分改变在诱发能量代谢紊乱和胰岛素抵抗，促进NAFLD发展至NASH中也起重要作用。本文旨在总结近年来在探索肠-肝轴和肠道菌群改变与肝脏炎症发生、发展关系方面的研究进展，为拟定NASH治疗的新策略提供科学依据。

1 肠-肝轴相互作用

肠道菌群对肝脏疾病的影响至少可追溯到80年前。Hoefert首先报道小肠细菌过度生长普遍出现于肝硬化患者，并与疾病的严重程度直接相关[1]。由于解剖结构和位置的特殊性，肝脏作为一级淋巴器官直接处理肠道血流中大量的细菌内毒素和代谢产物，通过胆盐等底物和肠-肝循环调节肠道功能、激素和免疫反应。同时，由于肝脏与肠道间的双向作用，肠道的激素、炎性介质和消化吸收产物也能调节肝功能。肝硬化时小肠动力障碍和口-盲肠传输时间延长是小肠细菌过度生长的生理基础，同时肠道黏膜通透性增加为肠道内细菌过度生长、侵入腹腔创造了条件，从而更易发生自发性细菌性腹膜炎和肝性脑病[2,3]。这是肠-肝轴影响肝脏疾病发生、发展的典型案例。

2 肠道菌群改变影响NAFLD的发生

在人类的消化道栖息着亿万种不同种类的微生物，其中大部分细菌种系定植在结肠，细菌总数为1013～1014个，菌体量> 1 kg。细菌的数量和类别保持动态恒定是维持肠道内环境稳定的基础[4]。人类宏基因组包括嵌入人体的基因组和伴生的微生物基因组。作为一个整体，微生物基因组数量是人类基因组数量的100余倍[5]。消化道在出生时是无菌的，出生后细菌开始定植并发展成稳定的菌群。婴儿肠道中的双歧杆菌是分解母乳中寡糖的关键菌群，能否建立稳定的菌群是婴儿正常消化吸收功能的重要前提。婴幼儿肠道菌群不稳定是发生腹泻和消化吸收障碍的原因之一[6]。成年后消化道内菌群的数量和类别保持相对稳定，但个体间差异巨大。消化道不同解剖部位的细菌种类和数量也不同，升结肠的细菌含量为1011个细胞/g，远端回肠为107～108个细胞/g，近端空回肠为102～103个细胞/g[7]。尽管肠道菌群的种系估计超过5 000种，但只有少部分菌种在数量上占主导地位[8,9]，如厚壁菌门约占60%、拟杆菌门约占15%、放线菌门约占15%、疣微菌门约占2%、变性菌门约占1%、甲烷杆菌目约占1%[10]。其中，拟杆菌门合成氢气，厚壁菌门合成各种可作为人体能量来源的短链脂肪酸(short chain fatty acid，SCFA)，包括丁酸等。此外，同一门类中的微生物功能可具有高度多样化[11]。

研究表明，超重和肥胖儿童粪便中双歧杆菌比例下降，金黄色葡萄球菌比例上升[12]。与健康人群相比，肥胖人群体内厚壁菌门比例上升，而拟杆菌门比例下降[13]。某些肠道菌群能更高效地从食物中摄取能量，促进肥胖和NAFLD的发生。Miele等对35例组织学证实的NAFLD患者研究后发现，患者肠道黏膜通透性增加，黏膜上皮细胞间紧密连接缺失，带状闭合蛋白1(zona occludens 1，ZO-1)表达下调。其中60%患者出现小肠细菌过度生长，且与肝脂肪变的严重程度显著相关[14]。动物实验表明，高脂饲料喂养的小鼠粪便中厚壁菌门比例升高，肠道内SCFA合成增加[15]。SCFA作为机体重要的能量来源，能增强肝脏自身脂质合成和三酰甘油聚集；同时也是肠上皮细胞、肠内分泌细胞和脂肪细胞的G蛋白偶联受体(G protein-coupled receptor，GPCR)的配体[16]。通过与GPCR结合，SCFA可增强胃肠激素如胰高血糖素样肽1(glucagon-like peptide 1，GLP-1)和酪酪肽(peptide YY，PYY)的分泌，直接或间接影响胰岛素和胰高血糖素的产生，调节食欲和食物的摄入[17]。

此外，动物实验发现部分高脂饲料喂养的小鼠(称为“应答”小鼠)除体重增加外，空腹血糖和胰岛素水平都升高，伴有单核细胞趋化因子1(monocyte chemoattractant protein 1，MCP-1)和肿瘤坏死因子α(tumor necrosis factor α，TNF-α)等促炎因子分泌增加。将“应答”小鼠和“非应答”小鼠的肠道菌群分别移植至无菌小鼠后，接受“应答”小鼠肠道菌群接种的受体小鼠出现了脂肪肝，并伴有转录因子固醇调节元件结合蛋白1c(sterol regulatory element binding protein 1c，SREBP1c)、糖类应答元件结合蛋白(carbohydrate response element binding protein，ChREBP)和乙酰辅酶A 羧化酶等脂质合成限速酶基因表达上调；而“非应答”受体小鼠则无显著变化。与“非应答”受体小鼠相比，“应答”受体小鼠粪便内厚壁菌门比例显著升高，表明肠道菌群的改变促进NAFLD发生[18]。但此结果能否在人体得到证实，仍需多样本临床对照研究验证。由于厚壁菌门被认为是“肥胖菌群”，且能在同一种系间接种传播，故有学者将由此类细菌引起的能量代谢异常、肥胖和NALFD定义为“感染性疾病”[19]。这一论点能否得到公认，有待商榷。

3 肠道菌群调控肝脏炎症和肝纤维化

肠道细菌的产物，如脂多糖(lipopolysaccharide，LPS)、脂多肽、DNA和RNA，具有潜在的肝毒性，能促进炎症发生[20]。这些细菌产物通过机体天然免疫系统的病原体相关分子模式(pathogen-associated molecular pattern，PAMP)和损伤相关分子模式(damage-associated molecular pattern，DAMP)激活肝细胞、Kupffer细胞和肝星状细胞(hepatic stellate cell，HSC)表面和细胞内的Toll样受体(Toll-like receptor，TLR)，启动相应的炎症反应[21]。不同的TLR具有相应的配体和特异的PAMP和DAMP[22]。TLR是进化高度保守的Ⅰ型跨膜糖蛋白，包含2个结构域：富含亮氨酸的重复序列和细胞内信号结构域——Toll/白细胞介素1受体(Toll/interleukin 1 receptor，TIR)结构域[23]。TLR下游的信号通路包括髓样分化因子88(myeloid differentiation factor 88，MyD88)依赖或MyD88非依赖2条通路。MyD88依赖的下游信号通路激活核因子κB(nuclear factor κB，NF-κB)，促进TNF-α、白细胞介素6(interleukin 6，IL-6)、IL-8和IL-12等促炎因子和γ干扰素(interferon γ，IFN-γ)、MCP-1等免疫相关基因和趋化因子的转录；MyD88非依赖的信号通路主要促进下游IFN-β的表达[24]。其中，TLR4能通过与配体革兰阴性杆菌胞壁成分LPS结合，同时以MyD88依赖和MyD88非依赖2种途径激活下游信号通路[25]。

在小鼠NAFLD模型中，将普通饲料换成高脂饲料后，小鼠肠道菌群中厚壁菌门比例升高，拟杆菌门比例下降。小鼠体重增加，空腹血糖和胰岛素水平升高，肝内三酰甘油和炎性分子聚集，门静脉中LPS水平显著升高[26]。Henao-Mejia等研究表明，IL-1细胞因子超家族成员IL-18能通过调节肠道菌群，在NAFLD等代谢性疾病进展中起重要作用。IL-1β和IL-18被炎性小体复合物激活后，才能形成有生物学活性的分子。炎性小体复合物由半胱氨酸天冬氨酸蛋白酶1(cysteinyl aspartate specific proteinase 1, caspase-1)和核苷酸结合寡聚化结构域样受体(nucleotide-binding oligomerization domain-like receptor，NLR)家族蛋白组成。NLRP3和NLRP6天然免疫缺陷小鼠失去产生IL-18的能力，肠道菌群谱中普雷沃菌科和紫单胞菌科比例增加，导致细菌产物由肠道转位至血循环和肝脏，激活TLR4和TLR9。该小鼠在喂食胆碱缺失饲料后极易发生NAFLD，喂食高脂饲料后极易发生代谢综合征[27]。

NAFLD患者小肠细菌过度生长、小肠黏膜通透性增加及紧密连接丢失引起的细菌转位导致血浆中LPS浓度升高，这些改变与NAFLD向NASH进展相关[14,28,29]。与轻度肝脂肪变患者相比，中、重度肝脂肪变患者的肠道黏膜通透性增加，小肠细菌过度生长的程度更甚[30]。NASH患者小肠细菌过度生长的严重程度与TLR4表达增强和IL-8生成密切相关[29]。肝组织炎症时，Kupffer细胞和HSC中TLR4表达水平升高。Kupffer细胞首先对LPS作出反应，产生炎性细胞因子、趋化因子和活性氧(reactive oxygen species，ROS)[31]，并通过分泌转化生长因子β(transforming growth factor β，TGF-β)等促纤维化因子激活HSC[32]。与健康人群相比，NASH患者小肠细菌过度生长引起血浆中LPS增多，经LPS-TLR4信号途径诱导TNF-α水平显著升高[33,34]。除通过Kupffer细胞间接激活HSC外，肠道来源的LPS可直接通过TLR4激活HSC。LPS通过下调TGF-β负向调节受体Bambi的功能，强化TGF-β对HSC的激活，活化的HSC分泌CC趋化因子配体2(CC chemokine ligand 2，CCL2)和CCL4等趋化因子，招募Kupffer细胞，后者再分泌TGF-β，继而促进肝纤维化进展[32]。如此，LPS所致肝内非实质细胞间相互作用使肝脏炎症和纤维化反应得以持续。

其他肠道细菌产物如细菌DNA，可通过DAMP模式影响NASH的进展和慢性化。细菌DNA富含CpG序列，与细胞内TLR9结合后，通过MyD88招募和激活下游NF-κB等信号分子，促进炎症分子表达[35]。此外，TLR9还通过干扰素调控因子7(interferon regulatory factor 7，IRF-7)促进IFN-α表达[36]。肝细胞、肝血窦内皮细胞、Kupffer细胞和HSC都可功能性表达TLR9[37,38]。小鼠NASH模型中，细菌DNA通过与Kupffer细胞内TLR9结合，以MyD88依赖途径促进IL-1β表达。IL-1β通过活化HSC，上调Ⅰ型前胶原和金属 蛋白酶组织抑制剂1(tissue inhibitor of metalloproteinase 1，TIMP-1)等促纤维化基因表达及下调Bambi表达，促进肝纤维化。TLR-9缺陷和IL-1受体缺陷小鼠与野生型小鼠相比，脂肪性肝炎和肝纤维化程度显著下降[39]。

4 肠-肝轴可作为防治NASH和肝纤维化的新策略

肥胖和高脂饮食可改变肠道菌群，因此可通过抗生素、益生菌和益生元等药物调节肠道菌群，改善肠道黏膜屏障，抑制小肠细菌过度生长，降低外周血和门静脉内毒素水平，以达到控制NASH向肝纤维化进展的目标。

动物实验表明，双歧杆菌可改善高脂饲料小鼠的葡萄糖稳态，降低小鼠体重和体脂含量，恢复葡萄糖介导的胰岛素分泌，并降低促炎性细胞因子(如TNF-α、IL-6等)和LPS水平[40]。给予ob/ob小鼠口服益生菌复合制剂VSL#3(唾液链球菌、嗜热链球菌、双歧杆菌和嗜酸乳酸杆菌等8种活性益生菌混合物)，可降低肝内脂质含量和肝脏炎症，改善肝脏的胰岛素抵抗[41]，并能通过下调NF-κB活性降低促炎性细胞因子分泌，以及降低TNF-α、诱导型一氧化氮合酶(inducible nitric oxide synthase，iNOS)和环氧酶2等脂质过氧化标记的表达[42]。此外，VSL#3还可通过减少平滑肌α肌动蛋白聚集、降低前胶原α1表达、刺激Bambi受体表达、控制HSC活化等机制，延缓胆碱缺乏饲料诱导的NASH纤维化进程[43]。尽管益生菌在动物模型中能缓解肝脏炎症和降低肝内脂肪聚积，但用其治疗NAFLD的临床试验结果却不甚理想。因NAFLD患者在停药4个月后肝内脂质含量显著增加，故VSL#3治疗NAFLD的临床试验被提前终止[44]。

口服肠道不吸收抗生素可调节肠道菌群，但由于无法特异性针对“有害”细菌，故备受争议。研究表明，给予高脂饲料诱导的肥胖小鼠口服万古霉素，可使厚壁菌门与拟杆菌门细菌比例显著下降，变形菌门细菌数量降低。在万古霉素干预期间，尽管进食同等热量的食物，小鼠体重较对照组降低，空腹血糖、血浆TNF-α及三酰甘油水平显著下降[45]。在果糖诱导的小鼠NASH模型中，口服不吸收抗生素可显著降低血浆和门静脉LPS水平及肝脏TNF-α表达，减缓肝细胞脂肪变和肝脏炎症损伤[43]。但迄今为止，尚未有研究肯定益生菌或抗生素能显著减缓NASH患者向肝纤维化及终末期肝病进展。

肠道菌群与NAFLD的关系强调了肠-肝轴相互作用对机体能量代谢的影响。鉴于一些肠道细菌的组分和产物与NAFLD进展至NASH相关，人们试图通过定性和定量改变肠道菌群以延缓肝脏疾病的进程。尽管动物模型和部分临床研究显示，益生菌对NASH患者的肝脏损伤有潜在的治疗作用，但迄今尚无大规模的临床随机对照试验来验证这一结果。肠道菌群的类型和功能鉴定、小肠细菌过度生长与NASH起始和进展的因果关系，仍有待进一步研究。同时，NASH由胰岛素抵抗、肝脂质毒性、氧化应激反应等多因素引起，且常为肥胖、糖尿病及代谢综合征等疾病的肝脏表现或伴生疾病。NASH发病机制的个体化差异较大，一种机制往往不能解释所有患者的发病原因，故其治疗更应趋于个体化[46]。

[1] Hoefert B. Über die Bakterienbefunde im Duodenalsaft von Gesunden und Kranken [J]. Zschr Klin Med, 1921, 92: 221-235.

[2] Madrid AM, Cumsille F, Defilippi C. Altered small bowel motility in patients with liver cirrhosis depends on severity of liver disease [J]. Dig Dis Sci, 1997, 42(4): 738-742.

[3] Madrid AM, Hurtado C, Venegas M, Cumsille F, Defilippi C. Long-term treatment with cisapride and antibiotics in liver cirrhosis: effect on small intestinal motility, bacterial overgrowth, and liver function [J]. Am J Gastroenterol, 2001, 96(4): 1251-1255.

[4] Nicholson JK, Holmes E, Wilson ID. Gut microorganisms, mammalian metabolism and personalized health care [J]. Nat Rev Microbiol, 2005, 3(5): 431-438.

[5] Neish AS. Microbes in gastrointestinal health and disease [J]. Gastroenterology, 2009, 136(1): 65-80.

[6] Chichlowski M, German JB, Lebrilla CB, Mills DA. The influence of milk oligosaccharides on microbiota of infants: opportunities for formulas [J]. Annu Rev Food Sci Technol, 2011, 2: 331-351.

[7] Zoetendal EG, Vaughan EE, de Vos WM. A microbial world within us [J]. Mol Microbiol, 2006, 59(6): 1639-1650.

[8] Zoetendal EG, Rajilic-Stojanovic M, de Vos WM. High-throughput diversity and functionality analysis of the gastrointestinal tract microbiota [J]. Gut, 2008, 57(11):1605-1615.

[9] Eckburg PB, Bik EM, Bernstein CN, Purdom E, Dethlefsen L, Sargent M, Gill SR, Nelson KE, Relman DA. Diversity of the human intestinal microbial flora [J]. Science, 2005, 308(5728):1635-1638.

[10] Ley RE, Knight R, Gordon JI. The human microbiome: eliminating the biomedical/environmental dichotomy in microbial ecology [J]. Environ Microbiol, 2007, 9(1):3-4.

[11] Rajili-StojanoviM, Smidt H, de Vos WM. Diversity of the human gastrointestinal tract microbiota revisited [J]. Environ Microbiol, 2007, 9 (9):2125-2136.

[12] Kalliomaki M, Collado MC, Salminen S, Isolauri E. Early differences in fecal microbiota composition in children may predict overweight [J]. Am J Clin Nutr, 2008, 87(3):534-538.

[13] Ley RE, Turnbaugh PJ, Klein S, Gordon JI. Microbial ecology: human gut microbes associated with obesity [J]. Nature, 2006, 444(7122):1022-1023.

[14] Miele L, Valenza V, La Torre G, Montalto M, Cammarota G, Ricci R, Mascianà R, Forgione A, Gabrieli ML, Perotti G, Vecchio FM, Rapaccini G, Gasbarrini G, Day CP, Grieco A. Increased intestinal permeability and tight junction alterations in nonalcoholic fatty liver disease [J]. Hepatology, 2009, 49(6):1877-1887.

[15] Murphy EF, Cotter PD, Healy S, Marques TM, O′Sullivan O, Fouhy F, Clarke SF, O′Toole PW, Quigley EM, Stanton C, Ross PR, O′Doherty RM, Shanahan F. Composition and energy harvesting capacity of the gut microbiota: relationship to diet, obesity and time in mouse models [J]. Gut, 2010, 59(12):1635-1642.

[16] Le Poul E, Loison C, Struyf S, Springael JY, Lannoy V, Decobecq ME, Brezillon S, Dupriez V, Vassart G, Van Damme J, Parmentier M, Detheux M. Functional characterization of human receptors for short chain fatty acids and their role in polymorphonuclear cell activation [J]. J Biol Chem, 2003, 278(28):25481-25489.

[17] Samuel BS, Shaito A, Motoike T, Rey FE, Backhed F, Manchester JK, Hammer RE, Williams SC, Crowley J, Yanagisawa M, Gordon JI. Effects of the gut microbiota on host adiposity are modulated by the short-chain fatty-acid binding G protein-coupled receptor, Gpr41 [J]. Proc Natl Acad Sci USA, 2008, 105(43):16767-16772.

[18] Le Roy T, Llopis M, Lepage P, Bruneau A, Rabot S, Bevilacqua C, Martin P, Philippe C, Walker F, Bado A, Perlemuter G, Cassard-Doulcier AM, Gérard P. Intestinal microbiota determines development of non-alcoholic fatty liver disease in mice [J]. Gut, 2013, 62(12): 1787-1794.

[19] Chassaing B, Etienne-Mesmin L, Gewirtz AT. Microbiota-liver axis in hepatic disease [J]. Hepatology, 2014, 59(1): 328-339.

[20] Seki E, Schnabl B. Role of innate immunity and the microbiota in liver fibrosis: crosstalk between the liver and gut [J]. J Physiol, 2012, 590(Pt 3):447-458.

[21] Schwabe RF, Seki E, Brenner DA. Toll-like receptor signaling in the liver [J]. Gastroenterology, 2006, 130(6):1886-1900.

[22] Alisi A, Carsetti R, Nobili V. Pathogen- or damage-associated molecular patterns during nonalcoholic fatty liver disease development [J]. Hepatology, 2011, 54(5):1500-1502.

[23] O′Neill LA, Bowie AG. The family of five: TIR-domain-containing adaptors in Toll-like receptor signalling [J]. Nat Rev Immunol, 2007, 7(5):353-364.

[24] Guo J, Friedman SL. Toll-like receptor 4 signaling in liver injury and hepatic fibrogenesis [J]. Fibrogenesis Tissue Repair, 2010, 3:21.

[25] Freudenberg MA, Tchaptchet S, Keck S, Fejer G, Huber M, Schütze N, Beutler B, Galanos C. Lipopolysaccharide sensing an important factor in the innate immune response to Gram-negative bacterial infections: benefits and hazards of LPS hypersensitivity [J]. Immunobiology, 2008, 213(3-4):193-203.

[26] Cani PD, Amar J, Iglesias MA, Poggi M, Knauf C, Bastelica D, Neyrinck AM, Fava F, Tuohy KM, Chabo C, Waget A, Delmée E, Cousin B, Sulpice T, Chamontin B, Ferrières J, Tanti JF, Gibson GR, Casteilla L, Delzenne NM, Alessi MC, Burcelin R. Metabolic endotoxemia initiates obesity and insulin resistance [J]. Diabetes, 2007, 56(7):1761-1772.

[27] Henao-Mejia J, Elinav E, Jin C, Hao L, Mehal WZ, Strowig T, Thaiss CA, Kau AL, Eisenbarth SC, Jurczak MJ, Camporez JP, Shulman GI, Gordon JI, Hoffman HM, Flavell RA. Inflammasome-mediated dysbiosis regulates progression of NAFLD and obesity [J]. Nature, 2012, 482(7384):179-185.

[28] Sabaté JM, Jou⊇t P, Harnois F, Mechler C, Msika S, Grossin M, Coffin B.High prevalence of small intestinal bacterial overgrowth in patients with morbid obesity: a contributor to severe hepatic steatosis [J]. Obes Surg, 2008, 18(4):371-377.

[29] Shanab AA, Scully P, Crosbie O, Buckley M, O′Mahony L, Shanahan F, Gazareen S, Murphy E, Quigley EM. Small intestinal bacterial overgrowth in nonalcoholic steatohepatitis: association with Toll-like receptor 4 expression and plasma levels of interleukin 8 [J]. Dig Dis Sci, 2011, 56(5):1524-1534.

[30] Thuy S, Ladurner R, Volynets V, Wagner S, Strahl S, Königsrainer A, Maier KP, Bischoff SC, Bergheim I. Nonalcoholic fatty liver disease in humans is associated with increased plasma endotoxin and plasminogen activator inhibitor 1 concentrations and with fructose intake [J]. J Nutr, 2008, 138(8):1452-1455.

[31] Baffy G. Kupffer cells in non-alcoholic fatty liver disease: the emerging view [J]. J Hepatol, 2009, 51(1):212-223.

[32] Seki E, De Minicis S, Osterreicher CH, Kluwe J, Osawa Y, Brenner DA, Schwabe RF. TLR4 enhances TGF-beta signaling and hepatic fibrosis [J]. Nat Med, 2007, 13(11):1324-1332.

[33] Wigg AJ, Roberts-Thomson IC, Dymock RB, McCarthy PJ, Grose RH, Cummins AG. The role of small intestinal bacterial overgrowth, intestinal permeability, endotoxaemia, and tumour necrosis factor alpha in the pathogenesis of non-alcoholic steatohepatitis [J]. Gut, 2001, 48(2):206-211.

[34] Rabelo F, Oliveira CP, Faintuch J, Mazo DF, Lima VM, Stefano JT, Barbeiro HV, Soriano FG, Alves VA, Carrilho FJ. Pro- and anti-inflammatory cytokines in steatosis and steatohepatitis [J]. Obes Surg, 2010, 20(7):906-912.

[35] Takeuchi O, Akira S. Pattern recognition receptors and inflammation [J]. Cell, 2010, 140(6):805-820.

[36] Takeda K, Akira S. TLR signaling pathways [J]. Semin Immunol, 2004, 16(1):3-9.

[37] Gäbele E, Mühlbauer M, Dorn C, Weiss TS, Froh M, Schnabl B, Wiest R, Schölmerich J, Obermeier F, Hellerbrand C. Role of TLR9 in hepatic stellate cells and experimental liver fibrosis [J]. Biochem Biophys Res Commun, 2008, 376(2):271-276.

[38] Martin-Armas M, Simon-Santamaria J, Pettersen I, Moens U, Smedsrød B, Sveinbjørnsson B. Toll-like receptor 9 (TLR9) is present in murine liver sinusoidal endothelial cells (LSECs) and mediates the effect of CpG-oligonucleotides [J]. J Hepatol, 2006, 44(5):939-946.

[39] Miura K, Kodama Y, Inokuchi S, Schnabl B, Aoyama T, Ohnishi H, Olefsky JM, Brenner DA, Seki E. Toll-like receptor 9 promotes steatohepatitis by induction of interleukin-1beta in mice [J]. Gastroenterology, 2010, 139(1):323-334.

[40] Cani PD, Neyrinck AM, Fava F, Knauf C, Burcelin RG, Tuohy KM, Gibson GR, Delzenne NM. Selective increases of bifidobacteria in gut microflora improve high-fat-diet-induced diabetes in mice through a mechanism associated with endotoxaemia [J]. Diabetologia, 2007, 50(11):2374-2383.

[41] Ma X, Hua J, Li Z. Probiotics improve high fat diet-induced hepatic steatosis and insulin resistance by increasing hepatic NKT cells [J]. J Hepatol, 2008, 49(5):821-830.

[42] Esposito E, Iacono A, Bianco G, Autore G, Cuzzocrea S, Vajro P, Canani RB, Calignano A, Raso GM, Meli R. Probiotics reduce the inflammatory response induced by a high-fat diet in the liver of young rats [J]. J Nutr, 2009, 139(5):905-911.

[43] Velayudham A, Dolganiuc A, Ellis M, Petrasek J, Kodys K, Mandrekar P, Szabo G. VSL#3 probiotic treatment attenuates fibrosis without changes in steatohepatitis in a diet-induced nonalcoholic steatohepatitis model in mice [J]. Hepatology, 2009, 49(3):989-997.

[44] Solga SF, Buckley G, Clark JM, Horska A, Diehl AM. The effect of a probiotic on hepatic steatosis [J]. J Clin Gastroenterol, 2008, 42(10):1117-1119.

[45] Murphy EF, Cotter PD, Hogan A, O′Sullivan O, Joyce A, Fouhy F, Clarke SF, Marques TM, O′Toole PW, Stanton C, Quigley EM, Daly C, Ross PR, O′Doherty RM, Shanahan F. Divergent metabolic outcomes arising from targeted manipulation of the gut microbiota in diet-induced obesity [J]. Gut, 2013, 62(2):220-226.

[46] Wu J. Coumarin: an alternative candidate for the treatment of non-alcoholic steatohepatitis [J]? Br J Nutr, 2013, 109(9):1542-1543.