Label-free plasma proteomics reveals the regulatory mechanism of Chaige Qingsan Decoction on complement and interferon pathways against influenza.

This study investigates the material basis and mechanism of Chaige Qingsan Decoction (CGQSD) in early influenza A virus-induced respiratory infections using plasma proteomics and enzyme-linked immunosorbent assay (ELISA) validation. Ultra-performance liquid chromatography coupled with Orbitrap high-resolution mass spectrometry (UPLC-Orbitrap HRMS) characterized the chemical profile of CGQSD. In the discovery cohort, 54 plasma samples from 22 influenza A patients (paired pre- and post-treatment) and 5 healthy controls underwent label-free quantitative proteomics. Patients received CGQSD monotherapy (n = 15), oseltamivir alone (n = 4), or combined drug therapy (n = 3). Key candidate proteins were validated via ELISA in an independent cohort (76 samples from 32 patients and 6 controls). A total of 1444 proteins were quantified after removing those with > 75% missing values, with high technical reproducibility (quality control [QC] r > 0.98). Compared with healthy controls, 263 differentially expressed proteins (DEPs) were identified. CGQSD modulated 178 DEPs enriched in complement/coagulation and interferon pathways. ELISA confirmed the significant regulation of C4BPB (P = 0.0003) and IFI6 (P = 0.016), consistent with proteomics.CGQSD mediates anti-influenza activity through a dual immunomodulatory mechanism centered on "Complement-Interferon Homeostasis," targeting C4BPB (a complement regulator) and IFI6 (an interferon-stimulated gene). CGQSD monotherapy fine-tunes complement activation via precise C4BPB downregulation; combination therapy buffers this effect and normalizes the interferon pathway by universally decreasing IFI6. Notably, combination therapy triggers extracellular matrix remodeling (e.g., COL1A upregulation), indicating enhanced tissue repair beyond antiviral effects. These findings provide a molecular foundation for CGQSD and advocate for its further development as a host-directed immunomodulatory therapy.