Abstract
Ethnopharmacological relevance
Xiongzhi Qufeng Zhitong Granule (XZQF) is a classic traditional Chinese medicine (TCM) formula widely applied in clinical migraine treatment. Yet, the complex pathogenic mechanism of migraine remains incompletely clarified, which restricts the standardized clinical application and modern industrial transformation of XZQF. Therefore, further exploring its therapeutic effects and underlying molecular regulatory mechanisms against migraine is pivotal to support its clinical optimization and popularization.Aim of the study
This research aims to assessing the protective efficacy of XZQF against reserpine-triggered migraine injury in mice, with a core focus on exploring its modulatory effects on the 5-HT1B/TPH2/CGRP signaling axis, so as to supplement the mechanistic evidence for XZQF's anti-migraine effect.Materials and methods
A reserpine-induced migraine mouse model was successfully established in this study. We then systematically evaluated the general behavioral observation, pain threshold detection, serum and tissue biochemical index quantification, and cerebral histopathological staining after XZQF intervention. Simultaneously, we integrated the multi-dimensional research strategies of network pharmacology, multiple molecular docking, and molecular dynamics (MD) simulation to unveil and verify the core candidate ingredients and therapeutic targets of XZQF used in treating migraine. Finally, immunohistochemistry (IHC), immunofluorescence (IF), and Western blot (WB) experiments were performed to verify the potential key signaling pathways regulated by XZQF.Results
In vivo pharmacological results showed that XZQF intervention effectively reversed the weight loss and pain hypersensitivity of migraine model mice. Meanwhile, XZQF significantly balanced the abnormal levels of inflammatory-, pain-, and neurovascular regulatory- mediators in mouse serum and brain tissues, and alleviated pathological damage of cerebral neurons. Network pharmacology analysis screened out senkyunolide F, senkyunolide E, ferulic acid etc. as the candidate key components of XZQF. We further revealed that signaling pathways of the calcium signaling, PI3K-Akt, and cAMP were the targeted pathways. Importantly, molecular docking and 100 ns MD simulation results confirmed that senkyunolide F and senkyunolide E could bind stably to the 5-HT1B protein, suggesting the potential functions of XZQF on modulating 5-HT system. Subsequent molecular biological verification further validated that XZQF treatment markedly upregulated 5-HT1B expression, while inhibiting the protein levels of TPH2, CGRP, TRPV1, c-Fos, COX-2, and key factors in PI3K and cAMP pathways.Conclusion
The anti-migraine effect of XZQF is a synergistic result in the forms of "multi-component -target -pathway" regulation. Mechanistically, by activating 5-HT1B expression and suppressing the TPH2/CGRP axis, as well as inhibiting the overactivation of PI3K and cAMP signaling cascades, XZQF exhibits a comprehensively migraine alleviative efficacy. Collectively, our study offers novel preclinical basics for understanding XZQF's mechanism in treating reserpine-induced migraine, and lays new insights into its clinical translational development.