修回日期: 2017-02-23
接受日期: 2017-02-27
在线出版日期: 2017-03-18
非酒精性脂肪性肝病(nonalcoholic fatty liver disease, NAFLD)是全世界最多见和最重要的慢性肝病, 由于成人和儿童肥胖流行率增加, 其发病率迅速增加, 发病率高达17%-33%, 且有逐年上升趋势. 其中1/3-1/2可能为非酒精性脂肪性肝炎(non-alcoholic steatohepatitis, NASH), 单纯性脂肪肝随访10-20年发展为肝硬化的概率为0.6%-3.0%, NASH随访10-15年肝硬化的发生率高达15%-25%. 发生肝硬化后每年1%病例发生肝细胞癌. 有关NAFLD的发病机制迄今尚不完全明了. 普遍认为年龄、性别、肥胖、胰岛素抵抗、细胞因子、基因多态性、肠道微生物是主要的发病机制. 因此如能全面深入的了解发病机制, 可为治疗提供依据. 近几年陆续报道用细胞因子或基因为靶点, 对NAFLD治疗取得一定疗效, 在当前NAFLD尚无特效治疗的情况下, 基因或靶向治疗对疾病的预后有可能产生深远的影响. 应当着进一步深入的研究.
核心提要: 非酒精性脂肪性肝病(non-alcoholic fatty liver disease, NAFLD)的重要性在于非酒精性脂肪性肝炎(non-alcoholic steatohepatitis, NASH)由于炎症可进展到肝纤维化、肝硬化, 以致发生肝细胞癌. NAFLD是当前世界上最常见的肝病, 值得注意的是尽管单纯性脂肪肝预后良好, 但近年越来越多证据表明, 他在代谢综合征及2型糖尿病的发生发展中起关键作用, 而且他也可向NASH演变, 因此摸清NAFLD发生机制, 寻求有效的治疗方法显得极其重要.
引文著录: 池肇春. 非酒精性脂肪性肝病发病机制研究进展与现状. 世界华人消化杂志 2017; 25(8): 670-683
Revised: February 23, 2017
Accepted: February 27, 2017
Published online: March 18, 2017
Non-alcoholic fatty liver disease (NAFLD) is the most common and important chronic liver disease in the world. As the prevalence of obesity increases in adults and children, the incidence of NAFLD has increased rapidly, reaching 17% to 33%. NAFLD is clinically divided into two forms: simple fatty liver (SFL) and non-alcoholic steatohepatitis (NASH), with NASH accounting for 1/3-1/2 of all NAFLD cases. The probability of developing cirrhosis is 0.6%-3.0% in patients with SFL for 10-20 years, and as high as 15%-25% in patients with NASH for 10-15 years. Approximately 1% of cirrhosis cases develop hepatocellular carcinoma each year. The pathogenesis of NAFLD is still not completely clear. It is generally believed that age, sex, obesity, insulin resistance, cytokines, gene polymorphism, and intestinal microflora are involved in the pathogenesis of NAFLD. An in-depth understanding of the pathogenesis of NAFLD can provide a basis for treatment of this disease. In recent years, cytokines or genes have been reported as targets for NAFLD treatment with appreciated effects. Since there is currently no specific treatment for NAFLD, targeted therapy may have a profound impact on the prognosis of the disease.
- Citation: Chi ZC. Pathogenesis of non-alcoholic fatty liver disease. Shijie Huaren Xiaohua Zazhi 2017; 25(8): 670-683
- URL: https://www.wjgnet.com/1009-3079/full/v25/i8/670.htm
- DOI: https://dx.doi.org/10.11569/wcjd.v25.i8.670
非酒精性脂肪性肝病(non-alcoholic fatty liver disease, NAFLD)是指除外酒精和其他明确的损肝因素所致的, 以弥漫性肝细胞大泡性脂肪变性为主要特征的临床病理综合征. 包括单纯性脂肪肝(simple fatty liver, SFL)和非酒精性脂肪性肝炎(non-alcoholic steatohepatitis, NASH)2种, NAFL在一定条件下可向NASH演变. 早期NASH: 无或轻度(F0-F1)纤维化; NASH: 显著(≥F2)纤维化或进展期(≥F3, 桥接)纤维化; 少数患者进展为肝硬化甚至肝细胞癌.
NAFLD呈全球化流行的趋势, 已成为发达国家和中国沿海地区慢性肝病的首要病因. SFL随访10-20年发展为肝硬化的概率为0.6%-3.0%, NASH随访10-15年肝硬化的发生率高达15%-25%, 其中30%-40%将会死于肝癌、肝功能衰竭和移植后复发[1-3]. NAFLD在无肥胖、无超重[体质量指数(body mass index, BMI)<25 kg/m2]有7%的人发生NAFLD, 而超体质量者28%, 两者有显著差异, 也说明不是发生NAFLD的唯一因素. 用超声诊断NAFLD在美国流行率为18.8%-30.2%[4,5]. 有关NASH的流行率亚洲为1.1%-2.2%, 美国为2.6%, 且流行率还有性别和种族上的差异[6].
NAFLD的发病与肥胖或超重有密切关系, NAFLD的检出率随BMI的增加而增高[7]. 多种代谢紊乱并存者, NAFLD发病率更高, 而且NASH引起肝硬化的可能性更大[8]. 我国成人超重率为22.8%, 肥胖率为7.1%[9]. 在今后10年内可能会显著增加. 肥胖也是小儿人群全球重要的健康问题. 儿童NAFLD亦与肥胖和糖尿病密切相关, 常累及青少年, 但亦可在5-10岁发病. NAFLD的发病机制至今还不完全明了, 可能与遗传易感性、基因、基因多态性、肥胖、胰岛素抵抗、细胞因子、肠菌生态失衡等多个因素共同作用所致.
NAFLD发病率及NASH的发生与种族、民族和家庭密切相关, 因此认为NAFLD是一种遗传-代谢应激相关性疾病. 近年来的研究表明, 免疫紊乱, 特别是固有的淋巴细胞(ILC)在肥胖和代谢紊乱中发挥调控作用. 其中, ILC2在NAFLD发病中有保护作用, 而ILC2亚群的缺失ILC或异常突变会促进慢性炎症和疾病的进展[10]. NAFLD的种族差异提示了在脂肪肝发病过程中存在遗传因素.
表观遗传学是研究不涉及DNA系列改变的基因表达和调控的可遗传的变化, 或者说是研究从基因演绎为表型的过程及其机制的一门遗传学分支, 近几年的研究提示他与许多疾病(包括肿瘤)的发生有关. 在NAFLD时无DNA系列改变, 通过遗传表型影响基因表达, 证实在NAFLD发病机制上是一个新视觉, 可逆的表观遗传在转录水平上发生改变, 且证明宿主和环境之间的一个表型连结. 近年的研究提出, 表观遗传学作用在NAFLD像是病理机制、生化标记和治疗的靶[10,11].
Apo-B基因位于人类第2号染色体短臂的23-24区, 全长34 kb, 含有28个内含子和外显子, 其基因结构有丰富的多态性. NAFLD时BMI、腰臀比、甘油三酯(triglyceride, TG)、总胆固醇和Apo-B均显著升高, 提示存在严重的脂质代谢紊乱, 并有种族差异.
Apo-B7673C/T多态性位点少见等位基因在NAFLD时分布异常, 可能其并非NAFLD的易感基因. 中性粒细胞衍生的载脂蛋白2经CXCR2(G蛋白偶联受体超家族成员)诱导通过中性-巨噬细胞途径, 介导NASH发生[12].
Apo-C3是一种水溶性蛋白质, 由79个氨基构成的糖蛋白, 由肝及小肠合成, 主要分布于血浆高密度脂蛋白(high density lipoprotein, HDL)、超低密度脂蛋白(very low-density lipoprotein, VLDL)和乳糜微粒(chylomicron, CM)中. Apo-C3在富含三酰甘油酯蛋白如VLDL、CM的代谢中起着重要作用, 由于脂肪肝是以血TG和脂肪酸的增加为主, 因此考虑Apo-C3与脂肪肝有一定的关系. 人Apo-C3位于染色体11q23.3, 长340q bp, 含4个外显子和3个内含子. Apo-C3 3175G/C多态性位点与NAFLD的关系及其作用途径尚待进一步研究. 但有报告Apo-C3的过度表达易于发生脂肪变性以及胰岛素抵抗(insulin resistance, IR)的发生[13].
ApoE是CM及VLDL的组成部分, 其含量与血清TG的水平呈负相关, E2、E3、E4构成了ApoE基因的多态性, 与肥胖和NAFLD的发生有关[14].
代表内脏的BMI和腰围与NAFLD的存在呈正相关, 尤其是内脏肥胖与NASH有密切关系. 人类肥胖相关基因有300多个, 除Y染色体外的所有染色体均分布有肥胖相关基因, 但与人类肥胖密切相关的基因多分布在2, 5, 10, 11和20号染色体上. 这类基因所编码的产物决定着脂肪沉积的形式、程度以及IR的发展, 在SFL、NASH和纤维化的发生发展中起重要作用. 过度表达11β羟基类固醇脱氢酶-1(11β hydroxysteroid dehydrogenase-1, 11β HSD-1)的转基因, 引起11β HSD-1活性增强是将无活性的肾上腺皮质激素可的松转化成有活性的氢化可的松, 糖皮质激素可增加脂肪细胞内脂肪生成和脂肪合成代谢, 可引起腹型和一系列代谢综合征的表现.
性腺功减退在NAFLD的发生上相关, 发现在男性由于性腺功能减退NAFLD的流行率增高, 包括性腺萎缩性腺功能减退症以及女性在经绝期后. 在这些情况下雌激素受体拮抗治疗或有Turner综合征, 雌激素在肝脂质平衡发挥关键作用. 鼠模型显示雌性激素降低和低浓度的血清睾丸酮是发生NAFLD的独立因子, 并将开发新的治疗NAFLD疾病的疗法[13]. 肠激素调节多种生物功能, 在NAFLD的发病机制上发挥作用. 他受食物摄取、体质量和IR的影响[15,16].
与IR相关的基因有过氧化物酶体增殖物激活受体γ2(peroxisome proliferator-activated receptor γ2, PPARγ2)基因Pro12A1α多态性, 他能提高胰岛素敏感性, 降低2型糖尿病(type 2 diabetes mellitus, T2DM)的敏感性, 在维护葡萄糖和脂质平衡上发挥关键作用[17]. 脂联素可减轻IR, 与NAFLD相关的AdiopQ基因多态性有rs266729(C>G)和rs2241766(T>G)2个位点. 有关rs266729G等位基因突变对NAFLD发病的影响研究报告不一, 多数研究认为可通过抑制脂肪酸和三酰甘油的分解代谢, 促进NAFLD的发生和发展[18]. FoxO1基因细胞是调节细胞氧化应激反应及细胞增殖、细胞凋亡、细胞自噬、代谢和免疫反应等多种生理作用的转录因子. FoxO1转录因子是Fox家族中Fox亚家族主要成员, 广泛表达于成人各级强器官中. 目前认为FoxO1为其活性形式, 定位于细胞核, 可调节糖脂代谢相关的多因表达[19]. IR使血清中游离脂肪酸(free fatty acid, FFA)增多, 当过量的FFA超出肝脏通过线粒体氧化反应及LDL形式排入血的代谢能力时, 将导致脂肪变性发生.
NAFLD常有脂肪酸氧化不足, 此时肝脏脂肪即可在肝脏沉积引起脂肪肝. 肝脏三酰甘油的合成、贮存转运、氧化的基因将会影响脂肪变的强度, 并影响NASH和肝硬化的进程.
肝内脂肪酸及酸主要来源于: (1)肝窦中CM的"残骸"; (2)循环中FFA; (3)肝内由乙酰辅酶A(NADPH)和H+合成, 硬脂酰基辅酶A脱氢酶将饱和FFA转化成单饱和FAA, 这在肝脏合成TG以及脂肪肝的形成均起关键作用. 固醇调节元件结合蛋白-1c(sterol regulatory element binding protein-1c, SREBP-1c)为转录因子 , 可增强自身的转录活性, 并上调多个脂肪酸合成酶(fatty acid synthase, FAS)生成相关酶(如ACC1/FAS/SCD1等)的基因表达, 从而增强FA和TG的合成. 在NAFLD时, 胰岛素对SREBP-1c表达异常增高[20]. 胆固醇代谢与NAFLD的发病机制和严重性密切相关. 胆固醇通过基因因子影响流动膜和蛋白功能膜, 并可展现蛋白反应, 且生成毒性oxysterol, 游离胆固醇作用于肝Kupffer细胞和星状细胞产生炎症细胞因子和胶原, 可损伤肝细胞和激活Kupffer细胞[21,22].
新近发现了与血脂代谢相关的遗传基因, 即TM6SF2, 其是一个异义E167K(rs58542926C/T)突变与NAFLD相关. TM6SF2在体内参与肝细胞脂肪代谢, 与VLDL分泌有关. TM6SF2 rs58542926 E167K突变导致核蛋白功能丧失, 引起TG过度积累, 导致NAFL发生[23]. KK/HIJ小鼠的研究[24]发现, 賴氨酸和大黄酸可降低肝细胞炎症和脂肪浸润, 对肝脏有保护作用. 可显著降低SOD和GSH-px的作用.
脂肪酸的氧化代谢在脂肪肝的发生和发展中有重要的作用. 酰基辅酶A氧化酶是β氧化系统的始动酶, 是脂肪氧化反应的关键酶, 当酰基辅酶A氧化酶基因缺陷可引起严重的NASH. 活性氧(reactive oxygen species, ROS)和脂质过氧化物生成增加, 后两者可直接造成细胞损伤, 亦可通过激活炎性反应, 引起NASH和肝纤维化[25].
PPARα在脂质代谢的动态平衡中起着重要作用. 主要作用为促进FA氧化分解, 减轻组织脂质沉积. PPARα是一种由配体激活的转录因子, 能启动一系列与脂质有关的酶和蛋白的基因转录, 主要包括脂酰辅酶A氧化酶、脂酰辅酶A合成酶、中链脂酰辅酶A脱氢酶、细胞色素450、微粒体CYP4A、脂肪酸ω羟化酶和脂肪酸结合蛋白等. ω可上调中链脂酰辅酶A脱氢酶和长链脂酰辅酶A脱氢酶的表达, 从而增强线粒体的β氧化[26]. PPARα还抑制Apo-C3, 增加VLDL分解[27].
氧化应激是指机体来自分子氧的游离基或细胞内ROS过度产生和/或抗氧化防御功能减弱, 造成组织细胞损伤的一种状态. 在人体抗氧化防御系统中, 有2种蛋白发挥重要作用, 一个是线粒体超氧化物歧化酶(superoxide dismutase, SOD), 他是体内最重要的抗氧化酶, 是对抗ROS第一道防线, 其基因多态性与NAFLD患者肝纤维化程度有关, 另一个是线粒体解偶联蛋白2(uncoupling proteins 2, UCP2), 也有抗氧化作用, 脂肪肝时UCP2表达上调, 对ROS生成有抵制作用. 当肝细胞氧化应激反应所产生的损害超过肝脏自身防御和时, 肝脏就可能会从SFL发展为NASH和肝纤维化[28,29].
随着氧化应激的产生和发展, Kupffer细胞和巨噬细胞的活化, 细胞因子和趋化因子的产生在NAFLD进展中发挥着核心作用[30,31]. 线粒体锰超氧化物歧化酶(Mn-superoxide dismutase, MnSOD)是线粒体消除自由基的关键酶, 当线粒体异常无法清除体内超氧阴离子, 使得肝细胞直接受到氧化损伤时, 就会导致NAFLD的发生. NAFLD的男性患者肝内MnSOD含量减少, 他可增加氧化应邀反应和肝病的进展[32].
NAFLD是代谢综合征组分, 且常是IR的最初迹象, 约98%的NAFLD表现胰岛素对抗.
NAFLD发病机制的Day二次打击学说一直被引用, 近年提出多次打击学说. IR是NAFLD的始动及中心环节. 关于IR、葡萄糖诱导的高胰岛素血症、血清三酰甘油水平升高和肝脂肪变性之间的关系有许多研究, 目前认为IR是通过脂解作用和高胰岛素血症2种机制引起[33,34].
已有研究表明preptin与IR相关性疾病的发生关系密切. preptin是新近发现的一种多肽类激素[35], NAFLD的多元回归分析表明, 腰围、FINS和HOMA-IR(稳定模型IR指数)是影响preptin水平的危险因素, 且preptin水平随着BMI增加而逐渐增高[36]. 有多囊卵巢综合征的NAFLD患者血清preptin显著升高, 且随着脂肪肝程度的加重而增高, 血清preptin水平与HOMA-IR、FINS等IR相关指标呈正相关[37].
肥胖、T2DM和高脂血症被认为是NAFLD的重要危险因子. 然而NAFLD是IR的结果还是NAFLD引起IR仍不完全清楚. 近年来也普遍关注脂肪肝与IR之间的"不相关"现象. 认为IR并不是发生NAFLD的必备条件. 此外肥胖鼠模型也观察到, 抑制肝TG合成可减少肝脂肪变, 但并不改变对胰岛素的敏感性, 因此, TG合成减少不一定与三酰甘油含量直接相关. 据报道, TG增高伴GGT、CRP增高而脂联素降低者发生IR的可能性大, 进展为糖尿病的风险也较高[38]. 然NAFLD是IR、脂质代谢紊乱、遗传背景、环境因子等引起的一个多基因疾病的过程[39].
脂联素是脂肪组织分泌最为丰富的一种激素, 具有胰岛素敏感性、抗炎症、抗动脉硬化、凋亡前作用和抗细胞增殖作用. 循环脂联素的水平主要取决于基因、饮食、生理活性和腹部脂肪. 近年研究发现脂联素与NAFLD的发生发展密切相关.
3.1.1 脂联素与肝星状细胞: 肝星状细胞(hepatic stellate cell, HSC)激活是产生肝纤维化的基础. 正常肝HSC处于静止状态, 位于肝窦周间隙和肝细胞隐窝内, 在损伤因素作用下, HSC发生增殖和表型转换, 活化的HSC能分泌大量的Ⅰ、Ⅲ型胶原反应的ECM, 同时通过对胶原酶等的抑制, 使胶原降解减慢, 最终导致ECM在肝内沉积. 脂联素通过胱冬肽酶途径诱导活化的HSC凋亡, 逆转HSC活化, 改变活化HSC的生理功能, 抵制活化HSC成纤维基因表达, 减少HSC增殖, 脂联素能够抑制HSC增殖和迁移, 促进HSC凋亡, 从而抑制了胶原形成和肝纤维化进程. 在NAFLD患者中血清脂联素水平下降越明显肝纤维化程度越严重[40]. 同时通过阻止Smad2核易位, 抑制转化生长因子β1和结缔组织生长因子的合成, 也可抑制HSC的激活, 脂联素还降低α-平滑肌肌动蛋白和核抗原表达. 脂联素介导AMP-激活蛋白激酶α通路涉及抗氧化、抗炎症和脂质代谢[41]. 肥胖是肝纤维化主要的风险因子和低脂联素密切相关. 脂联素调节水-甘油跨膜输送的蛋白通道, 改变肝HSC功能状态. 研究指出, 脂联素是HSC激活的有力抑制剂, 脂联素引起水-甘油跨膜输送的蛋白基因和蛋白在人星状细胞表达上调, 所以低血清脂联素水平可能是肥胖影响HSC功能状态的机制[42]. 肥胖患者显示脂联素缺乏和对抗, 结果导致肥胖相关肝纤维化发生[43]. 脂联素基因敲除小鼠中, 过表达脂联素可减弱HSC的增殖及促进其凋亡, 显著拮抗NASH相关肝纤维的进展[44].
3.1.2 脂联素多态性与NAFLD: 至目前为止已发现脂联素基因多种标记的单核苷酸, 包括rs182052、rs16861205、rs822396、rs7627128、rs1501299、rs2241767、rs3774261、rs266729、RS822393等. 现已确定rs2241767、rs1501299和rs3774261是个体易感和NAFLD进展的易感因子[45]. 单倍体分析指出, A-T-A单倍体是NAFLD的保护因子, 而G-G-A单倍体是NAFLD的风险因子, 包括rs2241767、rs1501299和rs3774261与NAFLD易感性之间相关[46,47]. rs266729(-11377G/C)和rs822393(-4522C/T)多态性发生NAFLD的风险增加[48].
脂联素基因多态性与冠心病发生有相关性, 脂联素266729G等位基因发生冠心病的风险增加. 脂联素基因多态性与幽门螺杆菌(Helicobacter pylori, H. pylori)并存时, 脂联素基因促进子-11391G/A, 细胞外SOD基因是高风险发生NAFLD. 在NAFLD的发病机制上基因型与H. pylori相互作用. 因此, 有效的预防NAFLD发生, 将考虑根除H. pylori或调控基因表达[49].
新近报道[50]指出, 高脂肪饮食的鼠实验证实, 补充HLF(山楂叶黄酮)可显著降低体质量、肝质量、肝/体质量比率, 改善血清参数和肝功能异常, 显著降低肝脂肪蓄积, 此外, HLF增加循环脂联素水平, 上调肝脂联素尤其是脂联素受体2的表达, 激活AMPK, 提出HLF改善肝脂肪变是脂联素/AMPK通路增强所致, 临床上可见NASH缓解.
3.1.3 脂联素与IR和T2DM: 脂联素对胰岛素有调节作用, 生理浓度的脂联素能明显增加肝脏、骨胳肌对胰岛素的敏感性, 增加脂肪酸的β氧化, 降低循环中的FFA, 阻止脂质在巨噬细胞中积聚, 抵制肝脏糖原异生及肝糖输出. 应用RT-PCR法检测NAFLD患者脂肪组织中脂联素mRNA的表达水平, 用稳态模型法计算IR指数, 探讨NAFLD患者IR与脂肪组织脂联素基因表达的关系. 结果发现IR与脂联素基因表达、血浆脂联浓度呈负相关、与血清三酰甘油呈正相关[51]. 噻唑烷二酮药物可增强脂联素的表达, 给予NASH患者胰岛素增效剂后, 可见纤维化减轻, 提示脂联素与胰岛素敏感性和纤维化的消长密切相关.
此外, 研究指出, 10%-20%的NAFLD患者进展到NASH. STK25是关键因子是异位脂肪沉积、葡萄糖和胰岛素体内平衡的关键调节. 鉴于STK在NASH发病机制上的多种作用, 因此, 将来研究开发STK25抑制剂来防治NASH具有重要意义[52].
瘦素是一个重要的脂肪因子, 由167个氨基酸组成的脂肪源性多肽激素, 人类瘦素主要由白色脂肪组织分泌, 棕色脂肪、骨骼肌、胃黏膜、胎盘、HSC等组织也可分泌. 机体的体脂肪量是影响瘦素分泌的主要因素, 尤其是腹部脂肪[53]. 促进瘦素分泌的因子尚有胰岛素、肾上腺素、TNF-α、IL-2β和大肠杆菌胞壁脂多糖(lipopolysaccharide, LPS), 而禁食、β-受体阻滞剂、环一磷酸腺苷(cyclic adenosine monophosphate, cAMP)、睾酮、生长激素则抑制其分泌.
瘦素能够促进HSC的纤维化发生反应以及增强金属蛋白酶组织抑制剂1的表达, 瘦素发挥促纤维化作用, 部分是通过抑制PPARγ, 他可逆转HSC的活化、维持HSC的静息表型. NASH时通过瘦素介导NADPH氧化酶、Micr-RA21作为调节TGF信号和纤维化的一个关键.
目前认为瘦素与胰岛素之间存在双向调节作用. 一方面瘦素通过对胰腺β细胞的作用而控制胰岛素的分泌, 而另一方面胰岛素又可刺激瘦素产生. 糖尿病前期患者瘦素和胰岛素指数测定, 研究[53]表明瘦素是NAFLD的风险因子 , 并发现有性别上的不同, 男性糖尿病前期NAFLD患者, 血清瘦素与空腹胰岛素、餐后胰岛素和HOMA-IR呈正相关, 与HOMA%S(胰岛素敏感性)呈负相关, 与BMI、TG和HOMA%B(胰岛素分泌功能)呈显著正相关.
研究[54-57]表明, NASH患者有瘦素或其受体基因变异, 因此不能正常发挥其调节肝脏对糖和脂肪代谢的功能, 致使肝细胞内抗氧化分子减少或活性降低, 导致线粒体内ROS过度生成, 而脂质过氧化引起NASH发生. 同时瘦素还参与NASH肝脏间质炎症反应, 致炎症反应加重. 另一方面炎症介质可选择性地使巨噬细胞分泌TNF-α、IL-6、IL-12, 也可促进炎症反应发生.
瘦素受体(leptin receptor, LEPR)多态性与脂质代谢和IR有关, 发现LEPR Q223RA等位基因显著降低, 是发生NAFLD和冠状动脉硬化的风险因子[55]. LEPR基因Gln223Arg(G/G)、MSOD9Ala/Val(V/V)和吸烟是NAFLD的风险因子[56]. 在鼠的试验指出, 二甲双胍像是一个减肥药, 可提高LEPR敏感性, 从而改善脂肪变[57], 瘦素治疗不仅减轻脂肪变, 可减少NASH发生[58], 而瘦素缺乏的肥胖小鼠限制蛋氨酸可防止脂肪变进展. 这是与蛋氨酸降低胰岛素和HOMA比率与Scd1基因下调有关[59]. 在肥胖青少年儿童低adropin是NAFLD独立危险因子, 评估血清adropin浓度可能是一个可靠的脂肪性肝病的标记[60].
总之, 瘦素对NAFLD有双向作用, 一方面瘦素可逆转IR和改善严重肝脂肪变性, 降低血清三酰甘油水平, 降低肝脏和肌肉组织内三酰甘油的比例, 提高机体对胰岛素的敏感性; 另一方面瘦素或其受体自身变异引起瘦素抵抗和高胰岛素血症, 形成高瘦素水平又可诱发IR.
固醇调节元件结合蛋白(sterol regulatory element binding protein, SREBP)是一类位于内质网上的膜连接蛋白, 是一种核转录因子属于"碱性螺旋-环-螺旋-亮氨酸拉链"超家族的一员. 迄今为止发现3种: 即SREBP-1a、SREBP-1c和SREBP-2. SREBP-1c主要参与脂肪酸的从头合成, SREBP-2促进胆固醇的合成, 而SREBP-1a则同时具有上述两种功能[61].
SREBP-1c是SREBP的一个亚型, 动物体内的SREBP-1 90%由SREBP-1c构成, 他是脂肪合成有关基因转录的决定因子, 故又称脂肪细胞决定和分化因子1(adipocyte determination and differentiation factor 1, ADD1), 主要在肝细胞和脂肪细胞表达. 他是参与脂肪合成基因的主要转录调节因子, 调节脂肪合成, 主要与脂肪酸和糖代谢有关[62]. 他调节的靶基因包括低密度脂蛋白(low density lipoprotein, LDL)受体、乙酰辅酶A羧化酶(ACC)、FAS、硬酯酰辅酶A去饱和酶(SCD)、葡萄糖激酶和磷酸烯醇式丙酮酸羧激酶等. SREBP-1c是介导胰岛素对葡萄糖激酶基因表达调控作用的转录因子. 基因敲除小鼠将会导致SREBP表达的减少, 最终导致肝脏胆固醇和脂肪酸的合成减少[63].
肌球蛋白轻链激酶(myosin light chain kinase, MLCK)是免疫球蛋白超家族蛋白质的成员. MLCK使内皮细胞上的肌球蛋白轻链(myosin light chain, MLC)磷酸化, 导致肠黏膜屏障功能减弱, 在NAFLD发病机制中发挥重要作用.
大量临床及动物实验研究结果显示NAFLD的疾病进展与肠黏膜机械屏障的受损有关. 肠黏膜机械屏障的损伤主要表现在肠黏膜通透性增高. Brun等[64]研究表明, NASH肥胖小鼠研究显示有肠道通透性增加和门静脉内毒素水平升高, 指出遗传性肥胖小鼠肠道通透性增加 , 导致了严重的门静脉内毒素血症. 提示NAFLD的发生与肠黏膜机械屏障的受损之间存在着紧密联系.
由此可见MLCK在肠道屏障功能形成障碍的机制中发挥核心作用. 由于MLCK表达增加, 肠黏膜机械屏障受损, 导致肠道通透性增加, 可促进《第二次打击》的危险因素更多更快的进入肝脏, 进而加速NAFLD的进展[65].
Ghrelin为新发现的生长激素促分泌素受体(growthhormone secreta-gogue receptor, GHSR), 是一种小分子多肽, Ghrelin与脂肪细胞GHSR结合而促进GH的分泌[66], 加速脂肪合成, 抑制脂肪分解, 致使体质量增加. 同时研究还发现低水平Ghrelin可诱发机体代谢紊乱, 导致肥胖、糖尿病等代谢疾病的发生[67].
Ghrelin对NAFLD似有一种保护作用. Cheyuo等[68]发现Ghrelin可能通过T淋巴细胞发挥抗炎作用. Ghrelin还可减少成肌纤维细胞及HSC聚集, 降低氧化应激和炎性反应程度, 减轻肝细胞损伤, 最终达到抑制肝纤维化的作用. 因此, 通过调节Ghrelin水平来干预NAFLD的发生发展将会成为一个新的治疗途径[69].
视黄醇结合4蛋白(retinol binding protein, RBP4)是一种新发现的脂肪因子, 不仅参与IR和脂质代谢紊乱, 还可引起炎症因子增高, 可能参与NASH进展. RBP4主要在肝实质细胞的粗面内质网中合成, 近年研究发现RBP4也可由脂肪细胞产生.
目前认为RBP4与IR和NASH进展相关: (1)脂肪分泌的RBP4在IR和NAFLD形成上发挥作用. PI3K是胰岛素刺激葡萄糖转运和摄取的重要介导分子, 胰岛素受体底物1(insulin receptor substrate, IRS1)酪氨酸磷酸化亦是胰岛素信号传导途径中的重要环节. RBP4直接诱导磷酸烯醇或丙酮酸激酶基因表达, 当PI3K活性下降和IRS1酪氨酸磷酸化水平降低时即会导致IR发. RBP4与肥胖、糖耐量受损和IR水平呈正相关[70,71]; (2)RBP4参与脂质代谢紊形成. 临床研究显示, 血清RBP4浓度g与TG、BMI、NAFLD明显正相关[72]; (3)RBP4可能参与NASH进展. RBP4增高可引起肝铁超载, SI和SF是评估肝铁超载的主要指标. 肝铁超载不仅加重了铁在肝脏中的沉积, 还可通过诱导炎症反应、氧化应激和抑制脂质转运, 从而导致NASH的发生与发展[73]. 但临床研究结果不尽相同, 有待进一步研究证实.
Toll样受体(toll-like-receptor, TLR)是一类模式识别受体家族, 可通过病原体相关的分子模式识别激活固有免疫系统发挥关键作用. TLR家族广泛参与机体免疫过程. TLR在肝脏的肝细胞、Kupffer细胞、肝窦内皮细胞、HSC和胆管内皮细胞等细胞中表达[74]. 大量研究[75,76]表明, TLR2、TLR4和TLR9 TLR也参与了NAFLD, 特别是NASH的发生与发展.
NAFLD时. Farhadi等[77]的研究提示, Kupffer细胞可与血液中高水平的TLR配体结合, 继而引发NASH患者的肝损伤. 在动物模型研究中, 发现肠源性内毒素和TLR在NASH发病中起到重要作用. Ye等[78]研究报告指出, TLR4在Kupffer细胞中的病理效应是通过诱导ROS依赖性活化的XBP-1(X盒结合蛋白)来实现的.
NAFLD患者循环中通常存在高水平FFA. 多项研究表明, FFA可通过TLR作用促进NAFLD发生和发展. LPS的脂质组分如中链脂肪酸、月桂酸可诱发巨噬细胞中的TLR4活化. Yamamoto等[79]研究认为棕榈酸可增强TLR2与其配体结合后的生理作用, 棕榈酸可能是TLR表达的启动因素, 通过激活Kupffer细胞/巨噬细胞引起炎症, 导致NASH进展. 新近Kim等[80]报告TLR7在NAFLD的发生上起到关键作用. TLR7信号途径被imiquimod(咪喹莫特, TLR7配体)激活, 通过诱导自噬和肝释放IGF-1可防止NAFLD进展, 提出NAFLD治疗新策略.
过氧化物酶体增殖物激活受体(peroxisome proliferator activated receptor, PPAR)是一类在能量代谢、脂肪细胞分化中起重要作用的核激素受体转录因子, 1990年从小鼠克隆获得, 因被过氧化物酶体增殖物激活后能诱导肝脏过氧化物酶体增殖而得名.
PPAR分α、β、γ 3种亚型, 分别由不同基因编码, 结构和功能各异. 其中PPARγ是脂肪细胞基因表达和胰岛素细胞信号间传递的主要调节者, 主要参与调节脂肪细胞的分化、脂肪酸的合成与贮存在. 可以减轻脂肪性肝炎, 抑制HSC激活和基质合成, 延缓脂肪肝纤维化的进程, 在NAFLD的发生、发展中具有重要作用. PPARγ有3种亚型, 即PPARγ1、PPARγ2和PPARγ3. PPARγ2特异性表达于脂肪组织中, 并可被高脂饮食所诱导, 也多含于HSC中.
PPARγ对脂肪细胞的增生和分化起着重要作用. PPARγ对脂肪细胞的分化起正向调节作用[80]. PPARγ除了在脂肪细胞分化中起关键作用外, 还在介导脂肪酸氧化及脂质代谢中起重要的作用. PPARγ表达升高诱导成脂性基因转录增多, 从而促进脂肪合成. PPARγ2基因C/G多态性与吸烟的协同效应通过增加氧化应激导致NAFLD的发展[81,82]. 研究结果表明, 在L-02肝细胞通过PPARα的抑制SREBP-1c表达上调FFA引起脂质蓄积和氧化应激[83].
NASH一般伴有IR, PPARγ激活物改善NASH的作用, 一方面主要通过脂肪细胞对胰岛素细胞敏感化过程起作用. 另一方面PPARγ可在体内、外抑制HSC的激活和基质合成, 导致肝纤维化的进程减慢. PPARα和PPARγ的双重激活通过一些肝和脂肪组织基因表达的调节对改善NASH有显著疗效[17]. PARP1是一种DNA损伤检测酶, 当这种酶抑制后, 可有效抵制癌症和其他疾病发生. 是一种可以检测并且对于DNA结构损伤进行信息反馈的蛋白. 在高脂肪饮食的鼠PARP1激活, PARP1药理或基因控制足以改变高脂肪包含引起的脂肪变和炎症. PPARα像是PARP1的底物, 介导PPARα的多聚ADP-核糖基抵制他聚集至靶基因启动子和他与Sirt1相互作用[84,85], 这是一个PPARα信号调节的关键, 导致脂肪酸氧化上调的抑制, 而且PARP1在人肝细胞是PPARα基因的一个转录抑制物, 且他的激活抑制配位体引起PPARα激活和靶基因表达. NAFLD患者的肝活检显示PARP活性和PPARα多聚ADP-核糖基水平增加. 研究结果提示, 在脂肪肝时PARP1激活, 通过PPARα信号的抑制以防止脂肪酸氧化的最大激活. PARP1的药理抵制可减轻PPARα的抵制, 因此, 对NAFL有治疗潜力[86].
微小RNA(microRNA, miRNA)是一类内生的、长度约为20-24个核苷酸的小RNA, 其在细胞内具有多种重要的调节作用, 每个miRNA可以有多个靶基因, 而几个miRNA也可调节同一个基因. 据推测, miRNA调节着人类三分之一基因. 随着研究的深入, 有关miRNA与NAFLD的报道也逐渐增多, 拓宽了对miRNA的认识, 对NAFLD的发病机制提出了新的见解.
直到目前为止与NAFLD相关的miRNA有miRNA-9、miRNA-124、miRNA-144、miRNA-33a、miRNA-301a-3p、miRNA-34a5p、miRNA-375等, miRNA表达是NAFLD进展的一个鲜明标志. miRNA-301a-3p、miRNA-34a5p表达增加, 并分析指出NAFLD严重性是伴有特殊类型肝miRNA表达, 改变脂质和碳水化合物代谢[87]. miRNA是转录后基因表达的调节者, 与NAFLD进展相关, 循环中miRNA可反映肝组织损害改变, 用于评价NAFLD的严重度[88]. miRNA-124通过TRB3促进肝TG蓄积[89]. miRNA-9与脂肪变之间呈正相关, Onecut2和SIRT1 2个miRNA-9的靶在NAFLD发生上发挥关键作用[90]. miRNA失调涉及各种肝的生化过程包括脂质体内稳定、炎症、凋亡和细胞增殖[88]. 新近许多文献报道miRNA与NAFLD进展有关, 因此, miRNA是评价NAFLD严重度的有力指标. , 为NAFLD的筛查和分期和监测疾病进展有重要价值[91].
miRNA在基因表达的微调上发挥重要作用, 他不仅是一个新的生化标记, 而且是NAFLD处理上的一个治疗工具. miRNA的失调与NAFLD R的不同期有关, 且与疾病的严重性相关[92]. Distefano等[93]也报告循环miRNA测定是在改善NAFLD的诊断和疾病进展的临床监测方法上一个新的发展.
肠道细菌对宿主起着免疫保护、营养物质的消化吸收、黏膜屏障、抗癌等多种作用[94]. 研究证实肠道细菌生态失衡也参与NAFLD的发生发展. 目前多项研究提示肠道菌群的改变可能是引起肥胖、代谢综合征的一个重要的环境因素[95]. 肠细菌生态失衡增加肠的渗透性和增加肝对损伤物质暴露, 增加肝炎症和纤维化, 如同时饮食调控不当, 短链脂肪酸(short-chain fatty acid, SCFA)和乙醇增加, 胆盐耗空, 细菌改变也可引起肠动力障碍, 肠道炎症和肠道免疫改变可导致肝损伤[96].
研究[97]发现NAFLD患者肠道通透性增高, 小肠细菌过度生长发生率增加. 饮食结构改变和抗菌素干预可调整肠道细菌, 为NAFLD的防治提出了新的挑战[98]. 当肠黏膜通透性增加时, 肠腔内大量细菌释放的内毒素经门静脉系统进入体循环, 形成内毒素血症[99], 后者可促使脂肪储存和IR发生. 另外, 细菌壁外膜上的LPS可通过TRL4作用于脂肪细胞和巨噬细胞, 诱导释放多种炎症细胞因子来诱发IR.
肠道细菌通过合成大量糖苷水解酶, 将植物多糖转变为单糖、SCFA、乙酸、丙酸、丁酸等. SCFA是大肠细菌代谢主要终产物, 主要由厌氧菌发酵难消化碳水化合物而产生. 肠道中的SCFA不仅可以作为营养物质而被吸收还可以影响机体脂类代谢和免疫反应等生物功能. 胰高血糖素样肽-1(glucagons-like peptide-1, GLP-1)是介导SCFA与肝脏脂肪代谢的重要物质, SCFA与存在于肠道内分泌细胞-L细胞膜表面的G蛋白偶联受体(G protein-coupled receptors, GPCR)41和GPCR43结合后, 通过增加细胞内的CA2+和cAMP浓度触发并加强L细胞分泌GLP-1与GLP-1受体结合, 达到控制血糖、降低体质量、降低血压、调节血脂、改善内皮细胞功能等多方面的代谢调节作用[100].
肠道细菌可通过胆汁酸的正常代谢间接对NAFLD的发生发展发挥作用. 正常时胆汁酸作为乳化液促进胃肠对脂肪与脂溶性维生素的吸收, 抑制肠道内菌群的过度繁殖和LSP释放, 以及控制肥胖等. 肠道细菌可通过法尼醇X受体(farnesoid X receptor, FXR)和G蛋白偶联胆汁酸受体(TGR)5调节胆汁酸的代谢, 并且参与有关胆汁酸合成、代谢和重吸收的基因表达. 饮食中的脂肪可改变胆汁酸的成分, 可显著地改变肠道细菌的组成并导致失调[101]. 胆汁酸还具有很强的杀菌作用, 通过与细菌细胞膜上的磷脂结合破坏菌膜, 达到抗细菌黏附并中和内毒素的效果, 抑制小肠细菌过度生长[102].
如上所述, 肠道细菌可通过FXR调节胆汁酸的代谢. FXR和他的下游靶点在调控肝脏脂质新生、HDL/TG输出和血浆TG转化中起到关键作用[103]. 使用TGR激动剂可降低血浆和肝脏TG水平, 从而减轻肝脏脂肪变性[104]. 因此, 通过调节胆汁酸的代谢和FXR/TGR5信号转导, 肠道细菌可直接促进NAFLD的进程[105,106].
益生菌在消化道腔内有黏膜防卫机制作用, 可限制病原菌定植, 把细菌黏连到黏膜表面, 阻止消化道细菌过度生长, 降低肠道菌群失调的发生率, 还可产生抗菌物质. 通过调节肠道菌群影响肠黏膜屏障的不同部分而提高肠道屏障功能, 对预防和延缓NAFLD的进展有重要作用[107]. 益生菌可减轻肝脏氧化应激和炎症损伤[108]. 益生菌治疗后, 不仅降低血清转氨酶水平, 还可改善IR[109]. 新近Sáez-Lara等[110] 人的临床试验报告, 益生菌可改善碳水化合物代谢、提高胰岛素敏感性, 改善IR, 降低血浆脂质水平, 使NAFLD和糖尿病获得好转. 目前临床上应用的益生菌种类繁多, 所含菌种不一, 因此, 需要扩大临床对照试验作进一步验证.
生活方式、饮食结构、性别和种族、肥胖、IR、遗传与基因多态性、激素和细胞因子、肠道菌群生态失衡等作为NAFLD的发病机制已得到共识, 尽管详细的机制尚未完全明了, 但目前已开始研究NAFLD的基因治疗和靶向治疗. 防治NAFLD不仅是医疗单位的事, 他需要全体民众的参与, 随着发病机制的进一步深入研究, 深信也必将给治疗带来新的曙光.
已往对非酒精性脂肪性肝病(non-alcoholic fatty liver disease, NAFLD)的发病机制一直沿用由Day倡导的二次打击学说, 近几年随着研究的深入提出多次打击学说, 当前认为性别、种族、遗传背景、细胞因子、基因多态性及基因受体、肠道菌群生态失衡等在NAFLD发生发展上有重要作用, 拓宽了对NAFLD发病机制的认识, 值得进一步研究.
有关NAFLD的发病机制的研究主要在动物模型中进行, 遗传背景、基因和细胞因子、生活方式和饮食结构、肠道菌群生态失衡是当前研究的热点, 但尚无重大突破, 随着认识的不断深入, 基因和靶向治疗的开展, 将会给治疗带来新模式.
通过对NAFLD发病机制认识的提高, 提出基因或靶向治疗新途径、新见解, 改善预后.
通过本文了解NAFLD发病机制的复杂性和多元性, 进一步认识肥胖、2型糖尿病、胰岛素抵抗及代谢综合征与NAFLD的相互关系, 为今后NAFLD的治疗提供理论依据.
姜春萌, 教授, 主任, 大连医科大学附属第二医院消化科; 涂相林, 主任医师, 南昌市第九医院(南昌大学附属感染病医院)
比较全面反映了NAFLD的发病机制研究现状, 展示了新的机制进展, 可为NAFLD的治疗新模式提供理论依据.
手稿来源: 邀请约稿
学科分类: 胃肠病学和肝病学
手稿来源地: 山东省
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