Immunomodulatory Effect of a New Ingredients Group Extracted from Astragalus Through Membrane Separation Technique

Abstract

Introduction: Astragalus is a commonly used traditional Chinese medicine in China, which has been widely applied to enhance the immunomodulatory function of the body. The main bioactive components are complicated. To explore the role of the components, various techniques have been applied in Astragalus extraction. Membrane separation technique featured with green processing condition and high efficiency is of signification interest in the application of Astragalus treatment.

Methods: In this study, a new ingredients group A4 was separated from Astragalus using membrane separation technique. The quantification and identification of A4 were achieved by UV-vis spectrometry and UPLC-MS measurements. Pathological approaches along with serum metabolomics were utilized to study the immunoprotective effects of the extracts and explore the underlying mechanisms on metabolic activity.

Results: It was observed that A4 could promote the secretion of IL-2 and IFN-γ, stimulate the activated CD4⁺CD25⁺ and CD8⁺ CD25⁺ T lymphocytes in splenocytes and protect rat spleen to some extent. Seven crucial biomarkers that related to immunity regulations were screened out and identified through serum metabonomic analysis coupled with nuclear magnetic resonance. The enrichment analysis revealed that A4 alleviated the immune dysfunction by modulating amino acid metabolism and energy metabolism for the first time.

Conclusion: The new ingredients group A4 isolated from the Astragalus membrane can reduce the immune dysfunction by regulating the amino acid metabolism and energy metabolism of rats.

Keywords: Astragalus; immunomodulatory; membrane separation; serum metabolomics.

Figure 1

Flow chart of Astragalus membrane separation process.

Figure 2

The content of (A) saponins and (B) flavonoids in AR stock and A4.

Figure 3

Pathological analysis of rat spleens (HE, ×200) in (A) control, (B) CTX, (C) CTX-A4, (D) HC, (E) HC-A4 groups.

Figure 4

Effects of A4 on the levels of IFN-γ (A) and IL-2 (B)in rats (±s, n=6). (**P<0.01 compared to the control group; ^ΔP<0.05 compared to the CTX group; ^#P<0.05 compared to the HC group).

Figure 5

Effects of A4 on CD4⁺CD25⁺ (A) and CD8⁺CD25⁺ T cells (B), as well as the ratio of CD4⁺CD25⁺/CD8⁺CD25⁺ (C) in rat splenocytes (±s, n=6) (**P<0.01 compared to the control group; ^ΔP<0.05 compared to the CTX group; ^#P<0.05 compared to the HC group).

Figure 6

PCA analysis from rat serum in CTX model (A) and HC model (B).

Figure 7

OPLS-DA analysis from rat serum in CTX model (A and B) and HC model (C and D).

Figure 8

Evaluation of each model using permutations substitution test. (A) Control vs CTX groups, (B) CTX vs CTX-A4 groups, (C) control vs HC groups, and (D) HC vs HC-A4 groups.

Figure 9

Bubble chart of metabolic pathways of A4 in (A) CTX-rats (1 serine and threonine metabolism; 2 glycerol metabolism; 3 glyoxylic acid and dicarboxylic acid metabolism; 4 glutathione metabolism; 5 pyruvate metabolism), and (B) HC-rats. (1 sweet starch and sucrose metabolism; 2 pyruvate metabolism; 3 galactose metabolism; 4 valine, leucine and isoleucine degradation; 5 valine, leucine and isoleucine biosynthesis).

Figure 10

A schematic diagram showing amino acids metabolism and energy metabolism interacting with each other during A4 treatment. The rectangles of different colors represent different metabolic pathways and the identified biomarkers were marked with star-shape.