# Mechanistic Exploration of Yiqi Zengmian in Regulating the Microenvironment as an Immunopotentiator with the Beijing Bio-Institute of Biological Products Coronavirus Vaccine Based on Transcriptomics and Integrated Serum Pharmacochemistry

## TL;DR

This study explores how Yiqi Zengmian boosts immunity when used with a coronavirus vaccine, revealing its active components and mechanisms.

## Contribution

The study identifies bioavailable components and validates the TLR-JAK-STAT pathway as a molecular target for YQZM’s immune-enhancing effects.

## Key findings

- YQZM significantly increased antibody titers, immune cell proportions, and cytokine levels in mice.
- 31 bioactive compounds in YQZM were identified using UPLC-MS/MS.
- The TLR-JAK-STAT pathway was confirmed as a key mechanism through RNA-Seq and molecular docking.

## Abstract

Background: Yiqi Zengmian (YQZM) functions as an immunopotentiator by enhancing both cellular and humoral immunity. However, its pharmacodynamic active constituents, particularly those absorbed into the bloodstream, and mechanism of action remain unclear. This study aimed to investigate the immunopotentiating effects and mechanisms of YQZM in mice immunized with the BBIBP-CorV (Beijing Bio-Institute of Biological Products Coronavirus Vaccine). Methods: Serum pharmacochemistry and ultra-performance liquid chromatography–tandem mass spectrometry (UPLC-MS/MS) were employed to identify bioavailable components of YQZM. The mice received the BBIBP-CorV twice on days 1 and 14, while YQZM was orally administered for 28 days. Neutralization assays and ELISA quantified antigen-specific antibodies (abs), flow cytometry (FC) and intracellular cytokine staining (ICS) were used to assess immune cell populations and their cytokines, and an enzyme-linked immunospot assay (ELISpot) quantified memory T and B cells (MBs and MTs). To identify underlying mechanisms, network pharmacology, RNA sequencing (RNA-Seq), molecular docking, Western blotting (WB), and quantitative reverse transcription PCR (RT-qPCR) were performed. Results: YQZM significantly enhanced antigen-specific antibody titers, immune cell proportions, cytokine levels, and memory lymphocyte functions. UPLC-MS/MS analysis identified 31 bioactive compounds in YQZM. KEGG enrichment analysis based on RNA-Seq and network pharmacology implicated the TLR-JAK-STAT signaling pathway in YQZM’s immune-enhancing effects. WB and RT-PCR validated that YQZM upregulated the expression of critical nodes in the TLR-JAK-STAT signaling pathway. Furthermore, molecular docking indicated that YQZM’s primary active components exhibited strong binding affinity for critical proteins. Conclusions: YQZM effectively enhances vaccine-induced innate and adaptive immunity via a multi-component, multi-target mechanism, among which the TLR-JAK-STAT signaling pathway is a validated molecular target.