Effect of electroacupuncture on hepatic autophagy in obese mice based on the AMPK/mTOR/ULK1 pathway

To investigate the effect of electroacupuncture (EA) on hepatic autophagy in obese mice by regulating the adenosine monophosphate-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR)/unc-51 like autophagy activating kinase 1 (ULK1) signaling pathway.

C57BL/6J mice were randomly divided into normal group, model group and EA group, with 8 mice in each group. The obese mice model was established by feeding high-fat diet. Mice in the EA group received EA at bilateral “Zusanli” (ST36) and “Tianshu” (ST25) for 30 min, 5 times a week for 4 consecutive weeks. The Lee’s index and body mass of mice in each group were observed before and after treatment; fasting serum glucose was measured; the insulin tolerance test (ITT) was performed, and the area under the curve (AUC) was calculated; the levels of fasting insulin, serum low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), total cholesterol (TC), and triacylglycerol (TG) were detected by ELISA, and the insulin resistance index was calculated; the mass of mesenteric white adipose tissue (mWAT) and liver tissue was weighed; the morphological changes of mWAT and liver tissue were observed by HE staining; the lipid deposition in the liver tissue was observed by Oil Red O staining; the ultrastructure of liver tissue was observed by transmission electron microscopy; the protein expressions of p-AMPK/AMPK, p-mTOR/mTOR, ULK1, microtubule-associated protein 1 light chain 3 type Ⅱ (LC3-Ⅱ), autophagy-related 5 (Atg5), autophagy-related 7 (Atg7), and sequestosome 1 (SQSTM1/p62) in liver tissue were detected by Western blot; the mRNA expressions of AMPK, mTOR, ULK1, LC3-Ⅱ, Atg5, Atg7, and p62 in liver tissue were detected by quantitative real-time PCR.

Compared with the normal group, the model group showed significant increases in Lee’s index, body mass, fasting blood glucose, fasting insulin, insulin resistance index, and ITT-AUC (P<0.01); increased serum TG, TC, and LDL-C levels (P<0.01); decreased serum HDL-C level (P<0.01); increased mWAT and liver mass (P< 0.01); enlarged adipocytes with reduced cell number per unit area in mWAT; swollen and disorganized hepatocytes with numerous lipid droplets and vacuoles; extensive orange-red lipid deposition; mitochondrial shrinkage and reduced autophagic vacuoles; decreased AMPK mRNA and p-AMPK/AMPK protein expression (P<0.01); increased mTOR mRNA and p-mTOR/mTOR protein expressions (P<0.01); decreased mRNA and protein expressions of ULK1, LC3-Ⅱ, Atg5, and Atg7 (P<0.01); and increased mRNA and protein expression of p62 (P<0.01). Compared with the model group, the EA group showed significant reductions in Lee’s index, body mass, fasting blood glucose, fasting insulin, insulin resistance index, and ITT-AUC (P<0.01); decreased serum TG, TC, and LDL-C levels (P<0.01); increased serum HDL-C level (P<0.01); reduced mWAT and liver mass (P<0.01, P<0.05); decreased adipocyte diameter and increased cell number per unit area in mWAT; no obvious lipid droplets or vacuoles in hepatocytes; reduced orange-red lipid deposition; normal mitochondrial structure, increased autophagic vacuoles, and occasional autophagosomes and autolysosomes; increased AMPK mRNA and p-AMPK/AMPK protein expressions (P<0.01); decreased mTOR mRNA and p-mTOR/mTOR protein expressions (P<0.01); increased mRNA and protein expressions of ULK1, LC3-Ⅱ, Atg5, and Atg7 (P<0.01); and decreased mRNA and protein expressions of p62 (P<0.01).

EA at ST36 and ST25 can promote hepatic autophagy in obese mice, and its mechanism may be related to the regulation of the AMPK/mTOR/ULK1 signaling pathway.