Astragalus mongholicus polysaccharides alleviate insulin resistance through modulation of PI3K/ AKT, TLR4/ NF-kB signaling pathway and microbiota in rats with Type 2 Diabetes Mellitus

Abstract

Background and aim: Astragali Radix has been widely used in traditional Chinese medicine to treat diabetes and a variety of other diseases. This study aims to evaluate the alleviating effects and mechanisms of Astragalus mongholicus Polysaccharide (mAPS) against diet combined with streptozotocin (STZ)-induced Type 2 Diabetes Mellitus (T2DM).

Experimental procedure: T2DM rats were orally administrated either with 200 mg/kg mAPS or 300 mg/kg Metformin (MET) once daily for four weeks. Body weight and Fasting Blood Glucose (FBG) were detected every 6 days. Serum fasting insulin (FINS) was measured by ELISA and the homeostatic model assessment of insulin resistance (HOMA-IR) was calculated accordingly. Histological change was studied by Hematoxylin and eosin (HE) staining. 16S rDNA sequencing was used to detect the changes in gut microbiota.

Results and conclusion: Oral administration of mAPS significantly decreased body weight, FBG, and HOMA-IR in T2DM rats (p＜0.05). Moreover, HE staining showed that mAPS could alleviate histological distortion in the liver and pancreas. Treatment with mAPS elevated the hepatic levels of phosphatidylinositol-3 kinase (PI3K), phospho-protein kinase B (AKT), and glucose transporter type 4 (GLUT4), while reducing phospho-nuclear factor kappa-B (NF-κB), Toll-like receptor 4 (TLR4), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-1β (IL-1β) (p＜0.05). Furthermore, mAPS supplementation could reverse the ratio of Firmicutes/Bacteroidetes (F/B) and reduce the abundance of Clostridia and Proteobacteria (p＜0.05). These results indicate that mAPS have the potential to enhance insulin sensitivity in diabetic rats by modifying gut microbiota and controlling the hepatic glycolipid metabolism and inflammation.

Keywords: Astragalus mongholicus polysaccharides; Gut microbiota; Insulin resistance; PI3K/AKT; T2DM.

Graphical abstract

Fig. 1

mAPS administration improved IR in T2DM rats. (A) Body weight. (B) FBG levels after 4 weeks of mAPS treatment. (C) Liver index. (D) Fat index. (E) HOMA-IR. (F) ISI. Results are expressed as the mean ± SD. Data with different letters are significantly different (p < 0.05).

Fig. 2

The effects of mAPS on serum metabolic parameters, liver and pancreas histopathology in T2DM rats. (A) TC, (B) TG, (C) ALT, (D) AST, (E) LDL, (F) HDL, (G) Representative HE staining of liver and pancreas (Original magnification: 200 × , scale bars = 50 μm), (H) Histological score of Liver, (I) Histological score of Pancreas. Results are expressed as the mean ± SD. Data with different letters are significantly different (p < 0.05).

Fig. 3

The effects of mAPS on the expression of tight junction proteins in T2DM rats. (A, B) The mRNA levels of the ZO-1 and Occludin. (C) The protein expression colon of ZO-1 and Occludin. Results are expressed as the mean ± SD. Data with different letters are significantly different (p < 0.05).

Fig. 4

The effect of mAPS on the hepatic PI3K/AKT pathways in T2DM rats. (A–C) The mRNA expression of hepatic PI3K, AKT and GLUT4. (D) The protein expression of hepatic IRS, PI3K, GLUT4, p-AKT, and AKT. Results are expressed as the mean ± SD. Data with different letters are significantly different (p < 0.05).

Fig. 5

The effects of mAPS on the hepatic TLR4/NF-κB pathway in T2DM rats. (A–B) Serum IL-1β and TNF-α levels. (C) The mRNA expression of hepatic IL-1β, TNF-α, NLRP3, TLR4, and IL-6. (D) The protein expression of hepatic TLR4, p-NF-κB, NF-κB, p-IκB, and IκB. Results are expressed as the mean ± SD. Data with different letters are significantly different (p < 0.05).

Fig. 6

Microbial diversity analysis in T2DM rats after mAPS intervention. (A) Venn diagrams. (B) α-diversity analysis of gut microbiota based on Shannon indexes. (C) β-diversity analysis of gut microbiota based on PCA.

Fig. 7

mAPS-regulated gut microbiota composition in T2DM rats. (A) Composition of bacteria phylum levels. (B) Composition of bacteria class levels. (C) The relative abundance of Firmicutes, Bacteroides, Proteobacteria, Clostridia. (D) The ratio of Firmicutes/Bacteroides. Results are expressed as the mean ± SD. Data with different letters are significantly different (p < 0.05).

Fig. 8

LEfSa analysis of gut microbiota composition in T2DM rats after mAPS treatment. (A) LEfSa multi-level classification tree diagram. (B) LDA-discriminant histogram (LDA >4).

Fig. 9

Correlation analysis between metabolic parameters and gut microbiota at the genus level. Note: *: p＜0.05，**: p＜0.01, ***: p＜0.001.

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