Natural exosomes-like nanoparticles in mung bean sprouts possesses anti-diabetic effects via activation of PI3K/Akt/GLUT4/GSK-3β signaling pathway

Abstract

Background: Type 2 diabetes mellitus (T2DM) is a chronic metabolic disease characterized by hyperglycemia and insulin resistance. Mung bean sprouts are traditionally considered a "folk" hypoglycemic food and their pharmacological effects and underlying mechanisms warrant further investigation.

Purpose: This study aimed to investigate the anti-diabetic effects of the exosomes-like nanoparticles in mung bean sprouts (MELNs) and explore the related molecular mechanisms.

Results: MELNs were isolated using a differential centrifugation-polyethylene glycol (PEG) method, and the identification of MELNs were confirmed by PAGE gel electrophoresis, agarose gel electrophoresis, thin-layer chromatography (TLC), and transmission electron microscopy (TEM). In the high-fat diet/streptozotocin (HFD/STZ) mouse model, MELNs ameliorated the progression of T2DM by increasing oral glucose tolerance test (OGTT) and insulin tolerance test (ITT) results, decreasing the fasting blood glucose level, and reducing the serum triglycerides (TG) and total cholesterol (TC). Histopathological examinations indicated MELNs diminished inflammatory infiltration of hepatocytes and amplified the area of islet B cells. In addition, MELNs decreased the oxidative stress levels in liver tissue and had good biocompatibility. In vitro experiments verified that MELNs improved the viability of glucosamine (GlcN) induced insulin-resistant hepatocytes. Furthermore, this study also revealed that MELNs upregulated GLUT4 & Nrf2 and down-regulated GSK-3β via activating the PI3K/Akt signaling pathway, promoting the production of antioxidant enzymes, such as HO-1 and SOD, to reduce oxidative stress.

Conclusion: MELNs mitigated the progression of type 2 diabetes in HFD/STZ mouse model. The underlying molecular mechanism is related to PI3K/Akt/GLUT4/GSK-3β signaling pathway.

Keywords: Exosome-like nanoparticles; Mung bean sprouts; Oxidative stress; Type 2 diabetes Mellitus.

Fig. 1

Characterization of MELNs. (A)-(B) The appearance of bean sprouts and MELNs. (C)-(D) Particle size of MELNs as well as Zeta potential. (E) TEM. (F) Effect of MELNs on blood cells. (G)-(I) Proteins, lipids, and nucleic acids in MELNs. (J)-(K) Uptake of MELNs by hepatocytes

Fig. 2

Progression of type 2 diabetes in HFD/STZ mice. (A) Changes in body weight in mice. (B) Changes in fasting blood glucose in mice at the third and fourth weeks. (C)-(D) Changes of OGTT and ITT in mice. (E) Serum levels of TG, TC as well as AST and ALT. **p < 0.01, *p < 0.05 compared to the HFD/STZ mice

Fig. 3

MELNs improved liver pathological changes, lipid accumulation and function. (A)-(B) Liver morphological changes and organ index, *p < 0.05, compared to the HFD/STZ mice. (C) TG, TC levels in liver tissue. (D)-(E) HE staining of the liver and pancreas was performed. (F) Function parameters of liver. **p < 0.01, *p < 0.05 compared to the HFD/STZ mice

Fig. 4

MELNs did not significantly reduce the viability of hepatocytes and reduced the oxidative stress level after Glcn-induced insulin resistance. (A)-(B) CCK8 assay was used to determine the effect of MELN on the cell viability of normal hepatocytes and GlcN (+) hepatocytes. (C)-(D) Parameters of hepatocyte function. (E)-(G) Changes of intracellular ROS in GlcN (+) hepatocytes treated with MELN. **p < 0.01, *p < 0.05 compared to the GlcN (+) hepatocytes

Fig. 5

Expression of oxidative stress-related proteins. (A)-(E) Western blotting assays showed that MELNs promoted Nrf2 nuclear translocation and increased HO-1 expression. **p < 0.01, *p < 0.05 compared to the GlcN (+) hepatocytes. (F)-(J) In vivo experiments also showed that the expression level of HO-1 and the ratio of Nrf2(N)/Nrf2(C) expression were increased after administration of MELNs. **p < 0.01, *p < 0.05 compared to the HFD/STZ mice

Fig. 6

Expression of PI3K/Akt in tissues. (A)(D) The expression of P-Akt/Akt and P-PI3K/Akt was detected by immunofluorescence. (B)-(C) Western blotting results showed that MELNs increased the phosphorylation of Akt and PI3K. (E)-(F) in the tissues. *p < 0.05 compared to the HFD/STZ mice

Fig. 7

Protein expression of PI3K/Akt/GSK-3β pathway. (A)-(E) Expression of PI3K/Akt/GSK-3β and their phosphorylated proteins in hepatocytes in vitro **p < 0.01, *p < 0.05 compared to the GlcN (+) hepatocytes. (F)-(I) The protein expression of P-GSK-3β/GSK-3β in liver tissue was detected. **p < 0.01, *p < 0.05 compared to the HFD/STZ mice

Fig. 8

Changes in glycogen accumulation in liver tissue as well as assessment of glucose uptake capacity. (A) Accumulation of hepatic glycogen in tissues. (B) Glycogen changes in hepatocytes after GlcN induction. (C)-(E) Uptake of 2-NBDG by hepatocytes. (F) Flow cytometry was used to determine the ability of hepatocytes to absorb 2-NBDG. **p < 0.01, *p < 0.05 compared to the GlcN (+) hepatocytes

Fig. 9

MELNs increased membrane translocation of GLUT4 and protein expression levels of GS and GLUT4. (A)-(B) MELNs increased the protein expression of GLUT4 and GS in hepatocytes in vitro. **p < 0.01, *p < 0.05 compared to the GlcN (+) hepatocytes. (C) Expression of GLUT4 at the cell membrane. (D)-(E) Expression of GS and GLUT4 in vivo. **p < 0.01, *p < 0.05 compared to the HFD/STZ mice

Fig. 10

The possible molecular signaling pathway for the exosomes-like nanoparticles in mung bean sprouts against diabetic effects