Abstract：
High-concentrate diet induce subacute ruminal acidosis (SARA) and cause liver damage in ruminants. It has been reported that forkhead box protein A2 (FOXA2) can enhance mitochondrial membrane potential but its function in mitochondrial dysfunction induced by high concentrate diets is still unknown. Therefore, the aim of this study was to elucidate the effect of high-concentrate (HC) diet on hepatic FOXA2 expression, mitochondrial unfolded protein response (UPRmt), mitochondrial dysfunction and oxidative stress. A total of 12 healthy mid-lactation Holstein cows were selected and randomized into 2 groups: the low concentrate (LC) diet group (concentrate:forage = 4:6) and HC diet group (concentrate:forage = 6:4). The trial lasted 21 d. The rumen fluid, blood and liver tissue were collected at the end of the experiment. The results showed that the rumen fluid pH level was reduced in the HC group and the pH was lower than 5.6 for more than 4 h/d, indicating that feeding HC diets successfully induced SARA in dairy cows. Both FOXA2 mRNA and protein abundance were significantly reduced in the liver of the HC group compared with the LC group. The activity of antioxidant enzymes (CAT, G6PDH, T-SOD, Cu/Zn SOD, Mn SOD) and mtDNA copy number in the liver tissue of the HC group decreased, while the level of H2O2 significantly increased, this increase was accompanied by a decrease in oxidative phosphorylation (OXPHOS). The balance of mitochondrial division and fusion was disrupted in the HC group, as evidenced by the decreased mRNA level of OPA1, MFN1, and MFN2 and increased mRNA level of Drp1, Fis1, and MFF. At the same time, HC diet downregulated the expression level of SIRT1, SIRT3, PGC-1α, TFAM, and Nrf 1 to inhibit mitochondrial biogenesis. The HC group induced UPRmt in liver tissue by upregulating the mRNA and protein levels of CLPP, LONP1, CHOP, Hsp10, and Hsp60. In addition, HC diet could increase the protein abundance of Bax, CytoC, Caspase 3 and Cleaved-Caspase 3, while decrease the protein abundance of Bcl-2 and the Bcl-2/Bax ratio. Overall, our study suggests that the decreased expression of FOXA2 may be related to UPRmt, mitochondrial dysfunction, oxidative stress, and apoptosis in the liver of dairy cows fed a high concentrate diet.
摘要：
高浓度饮食会引起反刍动物的亚急性瘤胃酸中毒（SARA）并引起肝脏损害。据报道，叉头盒蛋白A2（FOXA2）可以增强线粒体膜电位，但其在高浓缩饮食诱导的线粒体功能障碍中的功能仍未知。因此,这项研究的目的是阐明高浓缩（HC）饮食对肝脏FOXA2表达的影响，线粒体未折叠蛋白反应（UPRmt），线粒体功能障碍和氧化应激。选择总共12只健康的哺乳期中期荷斯坦奶牛，并随机分为2组：低浓缩物（LC）饮食组（浓缩物：饲草=4：6）和HC饮食组（浓缩物：饲草=6：4）。试验持续21天。瘤胃液，在实验结束时收集血液和肝组织。结果表明，HC组瘤胃液pH水平降低，pH低于5.6持续4h/d以上，表明饲喂HC日粮成功诱导奶牛SARA。与LC组相比，HC组的肝脏中FOXA2mRNA和蛋白质丰度均显着降低。抗氧化酶(CAT，G6PDH,T-SOD,Cu/ZnSOD,HC组肝组织中Mn一SOD)和mtDNA拷贝数降低，而H2O2的水平显著增加，这种增加伴随着氧化磷酸化(OXPHOS)的减少.HC组线粒体分裂和融合的平衡被破坏，OPA1，MFN1和MFN2的mRNA水平降低，Drp1，Fis1和MFF的mRNA水平升高。同时,HC饮食下调SIRT1、SIRT3、PGC-1α的表达水平，TFAM,和Nrf1抑制线粒体生物发生。HC组通过上调CLPP的mRNA和蛋白水平诱导肝组织UPRmt,LONP1，CHOP，Hsp10和Hsp60。此外,HC饮食可以增加Bax的蛋白质丰度，CytoC,Caspase3和Cleaved-Caspase3，同时降低Bcl-2的蛋白质丰度和Bcl-2/Bax比率。总的来说,我们的研究提示FOXA2的表达降低可能与UPRmt有关,线粒体功能障碍，氧化应激，高浓缩日粮奶牛肝脏的细胞凋亡。
