丁酸提高线粒体功能，减轻氧化应激，改善脂肪肝

1、畜禽脂肪肝

Fatty liver of livestock and poultry

脂肪肝问题在猪、鸡、牛、羊、鱼等各种动物中均普遍发生，成为影响动物健康和畜禽肉品质的重要因素，遗传、营养、管理、药物、毒素等均可导致脂肪肝的发生。脂肪肝的形成与脂肪代谢紊乱有关，肝细胞脂肪合成增加，氧化减少。氧化应激、NO信号通路的中断和线粒体功能障碍等被认为是加速脂肪变性和启动脂肪肝和纤维化进程的关键机制，并且线粒体损伤和氧化应激之间有着复杂的相互作用。

Fatty liver disease are widelly found in multiple animals, such as pigs, chickens, cattle, sheep, and fish etc., has become a vital factor to affect animal health and meat quality, and many factors such as genetics, nutrition, management, drugs and toxins etc. are possibly the reasons lead to a fatty liver. The formation of fatty liver is related to a disorder of the fat metabolism that the fatty synthesis of liver cells are increased and the oxidation are decreased. The oxidative stress, disruption of NO signaling pathway and mitochondrial dysfunction are considered to be the key mechanisms to accelerate steatosis and trigger the process of fatty liver and fibrosis’. And at the same, there also are some complicated interactions between mitochondrial damage and oxidative stress.

2、线粒体功能障碍

Mitochondrial dysfunction

线粒体有“细胞动力工厂”之称，除了为细胞提供能量，还参与细胞信号转导、分化与生长、凋亡等生命过程。作为肝细胞最重要的细胞器之一，线粒体是脂肪酸进行β-氧化和三羧酸循环、腺嘌呤核苷三磷酸（ATP）合成和活性氧（ROS）形成的主要场所。缺血缺氧、药物、毒素等都可导致线粒体功能障碍，表现为形态结构变化，ATP减少，游离氧产生过度，细胞凋亡、钙离子紊乱、mtDNA损伤等。

Mitochondria are known as the "Cell power plant". In addition to supplying energy to cellular, mitochondria are also involved in a range of processes, such  as  signaling, cellular differentiation and growth, and cell death. As one of the most important organelles of hepatocytes, mitochondria are the main site of fatty acid β-oxidation, tricarboxylic acid cycle, adenine nucleoside triphosphate (ATP) synthesis and ROS formation. Ischemia, hypoxia, drugs, and toxins etc. are the possibilties would lead to a mitochondrial dysfunction, which would manifeste as changes of morphological structure, ATP reduction, free oxygen’s excessive production, cell apoptosis, calcium disorder, and mtDNA damage, etc.

肝线粒体功能障碍可引起脂肪氧化的改变。线粒体脂肪酸β氧化是脂肪代谢的限速步骤，线粒体功能障碍导致肝细胞消耗游离脂肪酸的氧化磷酸化以及β-氧化减少，合成和摄取的甘油三酯增多，从而引起脂肪肝问题。

Liver mitochondria dysfunction causes some changes on fat oxidation. Mitochondrial fatty acid’s β oxidation is the rate-limiting step of fat metabolism, the mitochondrial dysfunction causes the oxidative phosphorylation of free fatty acids consumed by hepatocytes, the decreasing of β-oxidation, the increasing of the synthesization and ingestion of triglycerides, and that’s how the fatty liver is caused.

 肝线粒体功能障碍可引起活性氧(ROS)生成和氧化应激的改变。断奶、疾病、脂肪肝问题等在细胞水平上都是细胞的氧化应激，线粒体是氧化应激的作用靶点。已有大量研究报道在脂肪肝形成过程中，伴随着ROS的大量产生，线粒体氧化损伤产物如丙二醛（MDA）累积增加，线粒体内主要抗氧化蛋白GSH、SOD2和GPX 水平显著降低，抗氧化防御体系受到破坏，出现氧化应激，并进一步降低线粒体氧化呼吸功能。

The dysfunction of liver mitochondria will cause a production of ROS and change of oxidative stress. At the cellular level, weaning, disease and fatty liver problems are all the oxidative stress of cells and mitochondria is the target of oxidative stress. A large number of studies have reported that in the process of fatty liver formation, with the massive production of ROS, the accumulation of mitochondrial oxidative damage products such as malondialdehyde (MDA) starts to rise up, and the levels of the main antioxidant proteins GSH, SOD2 and GPX in the mitochondria are significantly reduced. The antioxidant defense system is damaged and the oxidative stress occurred, further reducing the mitochondria oxidative respiratory function.   

 

3、丁酸改善线粒体功能

Butyric acid improves mitochondrial function

丁酸作为一种重要的短链脂肪酸（SCFA），具有抗炎、抗癌、抗氧化和免疫调节等作用，既能作为能量基质直接被利用，也能作为信号分子调控基因和蛋白的表达。比如通过抑制组蛋白去乙酰化酶（HDAC）或激活G蛋白偶联受体41和43，来调控线粒体基因表达，影响机体代谢活动。

As an important short chain fatty acid (SCFA), butyric acid has the functions of anti-inflammatory, anti-cancer, anti-oxidation and immune regulation. Butyric acid can not only be directly used as the energy matrix, but also act as a signal molecule to regulate gene and protein expression. For example, by inhibiting histone deacetylase (HDAC) or activating G-protein-coupled receptors 41 and 43, butyric acid regulates mitochondrial gene expression and affects the body metabolic activity.

 丁酸可以通过减轻炎症反应、抑制胰岛素抵抗和减弱线粒体氧化应激等机制影响非酒精性脂肪肝的发生和发展。何进田等研究发现三丁酸甘油酯营养干预子宫内发育迟缓（IUGR）仔猪，可提高肝脏抗氧化能力，保护线粒体免受损伤；显著提高IUGR仔猪肝脏琥珀酸脱氢酶（SDH）、苹果酸脱氢酶（MDH）及锰超氧化物歧化酶（Mn-SOD）的活性，从而提高肝脏线粒体功能。

Butyric acid can affect the occurrence and development of nonalcoholic fatty liver by reducing inflammatory response, inhibiting insulin resistance and weakening mitochondrial oxidative stress. He Jintian et al. found out that tributyrin nutrition intervention in intrauterine growth retardation (IUGR) piglets will improve the liver antioxidant capacity, protect mitochondria from damage, and significantly improve the liver SDH, MDH and Mn-SOD activities of IUGR piglets, so as to improve the working performance of liver mitochondrial function.

                                  PS：SDH是三羧酸循环中唯一嵌入线粒体内膜的酶        MDH是一种重要的氧化还原酶        Mn-SOD主要存在于线粒体基质中，作为抗氧化剂PS: SDH is the only enzyme embedded in the inner mitochondrial membrane in the tricarboxylic acid cycleMDH is an important oxidoreductaseMn-SOD is mainly existed in the mitochondrial matrix as an antioxidant

 

丁酸可能通过增强肝线粒体功能缓解食源性小鼠肥胖，以及大鼠的非酒精性脂肪肝。Hatzis等研究表明，补充丁酸钠可以增强肠内褪黑素的合成，进而减弱内毒素诱导的活性氧的生成和肝脏氧化应激，从而对高脂饮食所诱导的肝脏疾病的保护作用。Mollica等也发现，丁酸盐能够通过激活AMPK/ACC通路，减少ROS生成，减弱氧化应激，调节线粒体的生物效率和功能状态，从而降低肝脏脂肪。The obesity in mice and nonalcoholic fatty liver in rats would be possibly alleviated by butyric acid through enhancing the function performance of liver mitochondria. Studies of Hatzis et al. showed that a supplement of sodium butyrate could enhance the synthesis of melatonin in the intestine, and by which have reduced the endotoxin-induced active oxygen production and liver oxidative stress, thereby to protect liver diseases induced by high-fat diet. Mollica et al. also found out that the butyrate could reduce the production of ROS, weaken the oxidative stress, and regulate mitochondrial biological efficiency and functional state, by activating AMPK/ACC pathway, thereby reducing liver fat.

 

 线粒体基因表达出现问题对能量代谢有着长期影响，丁酸可显著上调线粒体β氧化相关基因Acc1和Cpt1α的mRNA表达, 和解耦联相关的关键基因Ucp2的表达，以及线粒体自身编码的8个基因的mRNA水平的表达。

An expression trouble in the mitochondrial gene has a long-term effect on energy metabolism. The butyric acid can significantly up regulate the mRNA expression of mitochondrial β-oxidation related genes Acc1 and Cpt1α, and the expression of decoupling-related key genes Ucp2, as well as the mRNA level of eight genes encoded by mitochondria themselves.

 

qRT-PCR analysis for mRNA expression levels of the genes related with mitochondrial function in liver 

(A)The mRNA levels of mitochondrial function associated genes ACC1, CPT 1α and UCP2 

 

                                       
Con, normal diet；HF, high-fat diet；HFB, high-fat diet with sodium butyrate by gavagePS：PGC 1α是与机体能量代谢较为密切的转录辅助激活因子，在线粒体合成等过程中发挥重要作用。ACC1 和CPT-1α 是机体调控长链脂肪酸进入线粒体进行β氧化的重要酶。UCP2 在线粒体中与呼吸链电子传递和能量物质ATP的产生有重要的作用。PS: PGC 1α plays an important role in the process of mitochondrial synthesis.    ACC1 and CPT1α are important enzymes for regulating the β oxidation of long chain fatty acids into mitochondria.    UCP2 plays an important role in the electron transfer of respiratory chain and the production of ATP.

 

(B)The mRNA expression levels of 13 mtDNA-encoded genes

4、小结

Summary

畜禽脂肪肝的问题是能量摄入过多或者代谢异常，导致脂肪氧化不彻底以及氧化应激产生，根本原因是线粒体的功能不好，不能有效氧化脂肪释放出ATP，降低线粒体应激。用丁酸类衍生物干预后，线粒体功能改善，营养物质彻底氧化，ATP 的供应量充足，有助于减少脂肪在肝脏中的沉积，脂肪肝问题就解决了，动物也能好好地活下来。

The problems of fatty liver occured in livestock and poultry are caused on the account of an excessive energy intaken or an abnormal metabolism, and results in the incomplete fat oxidation and oxidative stress. The fundamental reason of this phenomenon is the poor function performance of mitochondria, which can not effectively oxidize fat to release ATP, and reduce mitochondrial stress. After an intervention of butyric acid derivatives, the mitochondrial function now has been improved, the energy substances is completely oxidized, the supply of ATP is sufficient, the problem of fatty liver is solved, and the animals is possible to survive well.

 从代谢动力学方面看，丁酸根在体内代谢速度特别快，在血液中6min达到峰值，很难维持有效的作用剂量和作用时间。而使用三丁酸甘油酯，血液中的丁酸根在15min达到效应浓度，然后持续升高，30min 时三丁酸甘油酯的含量达到峰值，丁酸根则在45min 达到峰值，并将效应浓度持续维持至使用后3 小时内，从而使足够量的三丁酸甘油酯和丁酸根能进入骨髓造血细胞里促进血红蛋白合成，提高氧气输入量，促进线粒体融合和再生，增强线粒体动力，输出更多的ATP，达到保命、促进肠道发育，降低脂肪肝等功效。

From the aspect of metabolism kinetics, butyrate root metabolizes very fast in vivo, with a 6-minute time to reach the peak value in blood, it is difficult to maintain an effective dosage and action time. But when tributyrin was used, the concentration of butyrate root in blood reached the effect concentration within 15 minutes, and then increased continuously. At the time of 30 minutes, the content of tributyrin reached the peak, while butyrate root at the time of 45 minutes, and maintained the effect concentration until 3 hours after being used. So that enough doses of tributyrin and butyrate is possible to enter into bone marrow hematopoietic cells to promote hemoglobin synthesis, increase oxygen input, promote mitochondrial fusion and regeneration, enhance mitochondrial power, output more ATP, achieve life-saving, promote intestinal development, reduce fatty liver and many other effects.

参考文献：

[1] 党文呈, 邓烽丞, 李兆龙, 等. 丁酸在非酒精性脂肪性肝病发生发展中的作用[J]. 临床肝胆病杂志, 2020, 36(4):915-918

[2] 洪健, 贾逸敏, 赵茹茜. 丁酸通过增强肝线粒体功能缓解高脂诱导的小鼠肥胖[J]. 中国生物化学与分子生物学报, 2017, 33( 12) : 1266 - 1273

[3] 何进田, 董丽, 白凯, 等.三丁酸甘油酯对宫内发育迟缓哺乳仔猪肝脏抗氧化和线粒体功能的影响亮.[J]. 食品科学, 2016, 37(3): 191-196

[4] Hatzis G, Ziakas P, Kavantzas N, et al, Melatonin attenuates high fat diet-induced fatty liver disease in rats[J]. World J Hepatol, 2013,5(4):160-169

[5] Jin C J, Engstler A J, Sellmann C, et al. Sodium butyrate protects mice from the development of the early signs of non-alcoholic fatty liver disease: Role of melatonin and lipid peroxidation[J]. Br J Nutr, 2016:1-12

[6] Mollica M P, Mattace Raso G, Cavaliere G, et al. Butyrate regulates liver mitochondrial function, efficiency, and dynamics in insulin-resistant obse mice[J]. Diabetes, 2017,66(5):1405-1418

[7] Wang Chunchun, Cao Shuting, Shen Zhuojun, et al. Effect of dietary tributyrin on intestinal mucosa development, mitochondrial function and AMPK-mTOR pathway in weaned pigs.[J]. Journal of Amimal Science and Biotechnology, 2019(1):1-10.