DOM FIGURE 4 Improved ATP homeostasis and attenuated ChREBP target gene induction after 4 week or 8 week treatment with AZD1656.

C57BL/6 mice were treated for either 4-wk (A-H) or 8-wk (I-P) without (0) or with AZD1656 (1 or 3mg/kg body wt) and used for the studies in Fig. 3E-H before they were euthanized for hepatocyte isolation. After overnight culture of hepatocytes, parallel incubations were performed in MEM with the substrates indicated (see legend to Fig. 2 for 4-wk treatments) for either 1h for determination of G6P, G3P, ATP (A-D,I-L) or 4h for mRNA analysis of the indicated genes (E-H,M-P). E,M: Basal mRNA levels at 5mM glucose normalized to vehicle 5mM glucose; F-H,N-P, mRNA levels of the indicated genes as % respective controls at 5mM glucose. Means ± SEM, n= 3-5 (4-wk, A-H) or n=4 (8-wk, I-P), *P< 0.05 vs respective control (5G); # P < 0.05 vs respective untreated (0 mg/kg).

本实验以C57BL/6小鼠为研究对象，分别接受4周（A-H组）或8周（I-P组）处理：未给药组予以0剂量处理，给药组予以AZD1656（1或3mg/kg体重）。于安乐处死分离肝细胞前，上述小鼠均用于图3E-H的相关实验。将分离得到的肝细胞过夜培养后，在含指定底物的最低必需培养基（Minimum Essential Medium, MEM）中开展平行孵育实验，其中4周处理组的底物信息详见图2图例；孵育时长分别设置为1h以检测葡萄糖-6-磷酸（glucose-6-phosphate, G6P）、甘油醛-3-磷酸（glyceraldehyde-3-phosphate, G3P）与三磷酸腺苷（adenosine triphosphate, ATP）（对应A-D、I-L组），或4h以对指定基因进行mRNA表达分析（对应E-H、M-P组）。其中，E、M组为5mM葡萄糖条件下的基础mRNA水平，以载体处理的5mM葡萄糖组为参照进行归一化；F-H、N-P组为指定基因的mRNA水平，以5mM葡萄糖条件下的对应对照组为基准，以百分比形式呈现。所有数据以平均值±标准误（SEM）表示，4周组（A-H）的样本量n=3~5，8周组（I-P）的样本量n=4；统计学显著性差异标注为：*P<0.05 与对应5mM葡萄糖对照组相比，#P<0.05 与未给药组（0mg/kg体重）相比。