Key Takeaways & Executive Findings
- •• Buyang Huanwu Tang improves motor function recovery in acute spinal cord injury rats, as evidenced by increased BBB scores and inclined plane test angles. • Proteomic and transcriptomic analyses identified key pathways and core targets, including complement and coagulation cascades, lysosome, and antigen processing, with FGG, FN1, FGB, and CXCL1 among the core differential proteins/genes. • The therapeutic mechanism involves inhibition of the Toll-like receptor 4/MyD88/NF-κB pathway to reduce inflammation, and regulation of BAX/BCL-2 balance to suppress Caspase-3 expression and apoptosis. • The study provides a comprehensive molecular basis for the neuroprotective effects of Buyang Huanwu Tang, supporting its clinical application in spinal cord injury.
Abstract
BACKGROUND: Research indicates that Buyang Huanwu Tang has positive therapeutic effects on the spinal cord injury symptoms and spinal cord function recovery, although its therapeutic mechanisms remain unclear. Spinal cord tissue contains numerous proteins and peptides that may serve as disease biomarkers. OBJECTIVE: To investigate the protective mechanism of Buyang Huanwu Tang in an acute spinal cord injury rat model by regulating proteomic- and transcriptomic-related pathways. METHODS: Thirty-six Sprague-Dawley rats were randomly divided into blank group, model group, and Buyang Huanwu Tang group. The latter two groups were used to establish acute spinal cord injury rat models using Allen's modified method. Motor function recovery was assessed by BBB score, pathological morphology was observed by Nissl staining, and differentially expressed proteins and genes were screened by isobaric tags for relative and absolute quantification (iTRAQ) proteomics and RNA-seq transcriptomics, followed by GO enrichment and KEGG pathway analyses. A PPI network was constructed using the STRING interaction database to identify key pathways, and core targets were validated by Western blot, immunohistochemistry, and RT-PCR. RESULTS AND CONCLUSION: (1) Motor function scores: Compared with the blank group, the model group showed significantly lower BBB scores (P < 0.001) and smaller inclined plane test angles (P < 0.001). Compared with the model group, the Buyang Huanwu Tang group showed higher BBB scores (P < 0.05) and larger inclined plane test angles (P < 0.001). Pathological morphology: The blank group showed relatively normal neuronal cells with intact structure, normal gaps, and clear nucleoli and nuclear membranes. The model group showed severe necrosis, disordered structure, pyknotic nuclei, disappearance of most nucleoli and nuclear membranes, numerous tissue cavities, and inflammatory cell infiltration. The Buyang Huanwu Tang group showed irregular but relatively intact neuronal cells with less swelling, reduced tissue cavities and cell necrosis. (2) GO functional annotation and KEGG pathway analysis revealed that differentially expressed proteins were mainly enriched in acute phase response, regulation of protein activation cascade, regulation of acute inflammatory response, platelet alpha granules, blood microparticles, vesicle lumen, serine-type endopeptidase inhibitor activity, and involved in pathways such as map04142 (lysosome), map04612 (antigen processing and presentation), map03013 (nucleocytoplasmic transport), map04964 (proximal tubule bicarbonate reclamation), map04610 (complement and coagulation cascades), map00511 (other glycan degradation), map03040 (spliceosome), map03410 (base excision repair), map00531 (glycosaminoglycan degradation), and coronavirus disease-COVID-19 pathway. Through PPI construction, 20 core differential proteins including FGG, FN1, FGB, HSP90B1, CASP3, and 20 core differential genes including FGG, FN1, FGB, CXCL1, CXCL13 were identified. (3) Western blot showed that Buyang Huanwu Tang inhibited the expression of myeloid differentiation factor 88 (MyD88) and P-IKBα in spinal cord tissue. Immunohistochemistry showed that Buyang Huanwu Tang inhibited the expression of BAX and Caspase-3, promoted BCL-2 expression, and inhibited apoptosis. RT-PCR results showed that Buyang Huanwu Tang inhibited the expression of MyD88 and CXCL1. These results suggest that Buyang Huanwu Tang may protect spinal cord tissue by inhibiting the Toll-like receptor 4/MyD88/nuclear factor-κB pathway to reduce inflammation, and by regulating the BAX/BCL-2 balance to inhibit Caspase-3 expression and prevent apoptosis.
1. Introduction
In recent years, the incidence of acute spinal cord injury (ASCI) has been increasing, becoming a major societal burden [1-3]. Buyang Huanwu Tang (BYHWT) has been widely used clinically due to its ability to inhibit inflammatory responses, improve ischemic conditions in spinal cord tissue, protect neural cells, and promote axonal regeneration [4-8]. However, the precise mechanisms underlying its therapeutic effects on spinal cord injury remain unclear, and its clinical application lacks effective theoretical guidance. Therefore, investigating the mechanisms of BYHWT has become a research priority.
Proteomics aims to explore the dynamic properties of all proteins within a specific context, including their structure, expression levels, post-translational modifications, interactions, and functions. Through these studies, proteomics enables qualitative and quantitative analysis of proteins, describing their dynamic changes and overall evolution patterns [9]. This approach helps identify objective biomarkers for ASCI and elucidate the intrinsic mechanisms of secondary injury. Furthermore, by analyzing changes in the proteome before and after BYHWT administration, it provides a possibility to explore the targets of traditional Chinese medicine and evaluate drug toxicity and efficacy. Currently, proteomic techniques mainly include two-dimensional electrophoresis, biological mass spectrometry, and liquid chromatography-mass spectrometry.
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Wang Ziqi, Bu Xianzhong, Guo Xiaohui, Li Hanxi, Qian Yuhao, Wang Yixin, Bu Baoxian (2026). Integrated proteomics and transcriptomics analysis of the mechanism of Buyang Huanwu Tang in protecting the acute spinal cord injury rat model. Chinese Journal of Tissue Engineering Research. https://doi.org/10.12307/2026.21524
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Frequently Asked Questions
What is the main objective of this study?
The main objective is to investigate the protective mechanism of Buyang Huanwu Tang in an acute spinal cord injury rat model by regulating proteomic- and transcriptomic-related pathways.
How was the acute spinal cord injury model established?
The acute spinal cord injury model was established using Allen's modified method in Sprague-Dawley rats.
What are the key findings of this study?
Buyang Huanwu Tang improves motor function recovery, reduces inflammation by inhibiting the Toll-like receptor 4/MyD88/NF-κB pathway, and prevents apoptosis by regulating BAX/BCL-2 balance and inhibiting Caspase-3 expression.
What techniques were used in this study?
The study used iTRAQ proteomics, RNA-seq transcriptomics, GO and KEGG pathway analyses, PPI network construction, Western blot, immunohistochemistry, and RT-PCR.
What are the core targets identified?
Core differential proteins include FGG, FN1, FGB, HSP90B1, CASP3, and core differential genes include FGG, FN1, FGB, CXCL1, CXCL13.
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