Juanbi Tang Attenuates Intervertebral Disc Degeneration by Suppressing p38MAPK/NF-κB Signaling and Activating Atg5-Dependent Autophagy to Promote Macrophage M2 Polarization: In Vivo and In Vitro Evidence.

backgroundIntervertebral disc degeneration (IDD) is driven by macrophage-mediated inflammation involving the p38MAPK/NF-κB signaling pathway and impaired autophagic flux. Juanbi Tang (JBT), a classical eleven-herb Chinese formula documented since 1732 CE for the treatment of Bi syndrome (a traditional diagnosis encompassing painful musculoskeletal obstruction conceptually corresponding to degenerative spinal conditions), demonstrates clinical anti-inflammatory activity; however, its molecular mechanism in IDD remains uncharacterized.

methodsJBT chemical composition was characterized by HPLC fingerprinting with osthole as quantitative reference standard (inter-batch RSD 2.3%) in accordance with the 2020 Chinese Pharmacopoeia, and quality control was extended beyond a single marker constituent by multi-component fingerprint similarity analysis using the cosine angle and correlation coefficient methods. A male Sprague-Dawley rat annular puncture IDD model (five groups: Sham, IDD, IDD + JBT, IDD + 3-methyladenine [3-MA], and IDD + JBT + 3-MA) was evaluated by MRI, CT, Safranin O-Fast Green, Masson trichrome and hematoxylin and eosin staining. LPS-stimulated (1 μg/mL) primary rat nucleus pulposus (NP) cells and murine RAW264.7 macrophages - including Atg5-overexpressing (Atg5-OE) and knockdown (Atg5-KD) variants - were used for in vitro mechanistic studies. Endpoints included apoptosis, mitochondrial membrane potential (ΔΨm), reactive oxygen species (ROS), macrophage M1/M2 polarization, ELISA cytokine quantification and Western blot pathway analysis.

resultsIn vivo, JBT significantly improved Pfirrmann grade, disc height index, Safranin O staining intensity, Masson-stained collagen content and Thompson grade, with concomitant suppression of serum TNF-α and restoration of IL-10; all benefits were partially abrogated by 3-MA co-administration. JBT suppressed p-p38MAPK and p-NF-κB p65 and restored the COL II/MMP-13 balance in disc tissue. In NP cells, JBT reduced total apoptosis from 29.1% to 14.5%, restored ΔΨm, attenuated ROS accumulation, reversed the Bax/Bcl-2 ratio, and activated autophagy (LC3-II ↑ , p62 ↓ ). In RAW264.7 macrophages, JBT promoted M2 polarization (CD206

conclusionsJBT attenuates IDD through coordinated suppression of macrophage-driven inflammation and activation of autophagy, providing mechanistic support for its clinical application in musculoskeletal disease.