Andrographolide attenuates neuroinflammation and apoptosis after spinal cord injury via PI3K-Akt pathway activation

Spinal cord injury (SCI) triggers secondary injury cascades dominated by microglia-driven neuroinflammation, mitochondrial dysfunction, and neuronal apoptosis, leading to progressive tissue loss and persistent neurological deficits. Andrographolide, a diterpenoid lactone from Andrographis paniculata, has broad anti-inflammatory activity, but its therapeutic potential and mechanism in SCI remain unclear. Here, we combined network pharmacology with experimental validation to evaluate andrographolide after SCI. Candidate targets and pathways were screened in silico, prioritizing PI3K-Akt signaling. In a mouse contusion SCI model, andrographolide was administered intraperitoneally at 30 or 60 mg/kg starting 2 h after injury and then once daily for 14 consecutive days. PI3K-Akt pathway involvement was examined using intrathecal LY294002 under the 60 mg/kg andrographolide regimen. Behavioral assessments were conducted from 0 to 28 days post-injury to evaluate the sustained effects of early andrographolide treatment. In vitro, LPS-stimulated BV2 microglia were treated with andrographolide with or without LY294002, followed by cytokine assays, JC-1 analysis of mitochondrial membrane potential, and p-PI3K immunofluorescence. Network pharmacology identified overlapping genes between andrographolide-related targets and SCI-associated genes, and highlighted PI3K-Akt as a key enriched pathway. Consistent with this prediction, andrographolide dose-dependently restored PI3K/Akt/GSK3β/CREB phosphorylation and shifted apoptosis markers toward survival in vivo, with 60 mg/kg producing stronger effects, while LY294002 attenuated these molecular benefits. Using the selected regimen, andrographolide improved locomotor recovery and gait performance, reduced lesion cavitation, and preserved neuronal survival after SCI. In BV2 microglia, andrographolide increased p-PI3K, suppressed LPS-induced TNF-α/IL-1β/IL-6 release, and restored mitochondrial membrane potential, whereas LY294002 attenuated these effects. Collectively, andrographolide confers neuroprotection after SCI in association with activation of PI3K-Akt signaling, attenuation of inflammatory responses, preservation of mitochondrial function, and inhibition of apoptosis. Complementary BV2 cell experiments suggest that microglia-related inflammatory modulation may contribute to these protective effects.