Artemether regulates liver glycogen and lipid utilization through mitochondrial pyruvate oxidation in db/db mice

Abstract

Objectives: Diabetes is an important global health problem. The occurrence and development of type 2 diabetes (T2D) involves multiple organs, among which the liver is an important organ. Artemether is a methyl ether derivative of artemisinin and has displayed significant antidiabetic effects. However, its regulation of glucose metabolism is not clearly elucidated. This study explored the effect of artemether on liver mitochondrial pyruvate metabolism.

Methods: T2D db/db mice were used and grouped into db/db and db/db+Art groups. Lean wild type mice served as control. After artemether intervention for 12 weeks, the respiratory exchange ratio (RER), redox state, relevant serum lipid content, liver glycogen and lipid content, liver insulin and insulin-like growth factor 1 (IGF-1) signal transduction, mitochondrial pyruvate oxidation pathway, fatty acid and glycogen metabolic pathways were evaluated.

Results: This experiment demonstrated that artemether raised RER and enhanced liver mitochondrial pyruvate metabolism in db/db mice. Artemether also reduced serum and urinary lipid peroxidation products and regulated the redox status in liver. The accumulation of liver glycogen in diabetic mice was attenuated, the proportion of lipid content in serum and liver was changed by artemether. The signal pathway associated with liver glycogen metabolism was also regulated by artemether. In addition, artemether increased serum insulin and regulated insulin/IGF-1 signal pathway in liver.

Conclusions: The present study confirmed that artemether can regulate liver glycogen and lipid utilization in T2D mice, its biological mechanisms were associated with mitochondrial pyruvate oxidation in the liver.

Keywords: Artemether; glycogen; lipid; liver; pyruvate.

Figure 1

Effect of artemether on RER and the major proteins involved in pyruvate oxidation in liver. A, B. RER values for the dark and light phase in each group. n=4 per group. C. Immunoblotting bands of p-PDH, PDH, PDK1, PDK4, MPC1, and MPC2 in various groups. D-I. Bar graphs displaying the fold changes of p-PDH, PDH, PDK1, PDK4, MPC1, and MPC2 in different groups. n=4 per group. *P<0.05, **P<0.01 and ***P<0.001 vs. control group; #P<0.05, ##P<0.01 and ###P<0.001 vs. the db/db group. J. Representative immunohistochemical staining images of PDK1 from each group. Scale bar, 100 µm.

Figure 2

Artemether reduced serum and urinary lipid peroxidation products, regulated the redox status in liver. A. The serum level of MDA in each group. B, C. The urinary 4HNE and 8-OHdG excretion in different groups. D. Bar graph showed the liver GSH content in each group. n=6 per group. E. Immunoblotting bands of SOD1, SOD2, SOD3, Catalase, GPX1, and GPX4 in various groups. F-K. Bar graphs displaying the fold changes of SOD1, SOD2, SOD3, Catalase, GPX1, and GPX4 in different groups. n=4 per group. *P<0.05, **P<0.01 and ***P<0.001 vs. control group; ##P<0.01 and ###P<0.001 vs. the db/db group.

Figure 3

Effects of artemether on fatty acid oxidation in liver. A. Immunoblotting bands of CPT1A, CPT2, CROT, CRAT, ACADVL, ACADL, ACADM, and ACADS in various groups. B-I. Bar graphs displaying the fold changes of CPT1A, CPT2, CROT, CRAT, ACADVL, ACADL, ACADM, and ACADS in different groups. n=4 per group. *P<0.05 and **P<0.01 vs. control group.

Figure 4

Artemether regulated signaling pathways associated with glycogen metabolism. A. Immunoblotting bands of p-GS, GS, PYGL, PCK1, PCK2, and G6Pase in various groups. B-G. Bar graphs displaying the fold changes of p-GS, GS, PYGL, PCK1, PCK2, and G6Pase in different groups. n=4 per group. **P<0.01 and ***P<0.001 vs. control group; ##P<0.01 vs. the db/db group.

Figure 5

Effects of artemether on serum and liver lipid contents. A-C. The serum levels of TG, FFA, and TC in various groups. D-F. Bar graphs displayed the liver contents of TG, FFA, and TC in different groups. G. Bar graph showed the lipid droplets area ratio in each group under HE staining. n=6 per group. *P<0.05, **P<0.01 and ***P<0.001 vs. control group; ##P<0.01 and ###P<0.001 vs. the db/db group. H. Representative images of HE staining of liver sections from each group. Scale bar, 100 µm.

Figure 6

Effects of artemether on liver weight and glycogen content in db/db mice. (A, B) Bar graphs display the liver weight and glycogen content in different groups. n=6 per group. **P<0.01 and ***P<0.001 vs. control group; ###P<0.001 vs. the db/db group. (C, D) Representative images of (C) PAS and (D) D-PAS staining of liver sections from each group (scale bar, 100 μm).

Figure 7

Artemether improved the insulin and IGF-1 associated signal transduction. A, B. The serum levels of insulin and IGF-1 in different groups. n=6 per group. C. Immunoblotting bands of IRS1 and IRS2 in various groups. D, E. Bar graphs displaying the fold changes of IRS1 and IRS2 in different groups. n=4 per group. *P<0.05, **P<0.01 and ***P<0.001 vs. control group; ##P<0.01 and ###P<0.001 vs. the db/db group. F. Representative immunohistochemical staining images of IRS1 from each group. Scale bar, 100 µm.