Combined transcriptomics and network pharmacology to elucidate the mechanisms of rutin in treating ischemic stroke rat

Background: Ischemic stroke (IS) is a neurovascular disorder characterized by a prolonged treatment period and a high risk of recurrence. Rutin, a principal flavonol, has demonstrated potential neuroprotective effects in previous studies; however, its regulatory mechanism on neuroinflammation following IS remains incompletely elucidated.

Purpose: This investigation aims to elucidate the neuroprotective mechanisms of rutin in IS, with a particular focus on the NF-κB/NLRP3 signaling pathway.: METHODS: A dual investigative strategy was employed, combining a transient middle cerebral artery occlusion (tMCAO)-induced cerebral ischemia model in vivo with oxygen-glucose deprivation and reoxygenation (OGD/R)-treated astrocyte cultures in vitro. These experimental approaches were integrated with network pharmacology prediction and transcriptomic profiling to systematically elucidate the neuroprotective efficacy and multitarget regulatory mechanisms of rutin.

Results: Our research demonstrated that rutin significantly reduces infarct volume and improves neurological outcomes in the tMCAO rat model, with the most pronounced effects occurring at a dosage of 80 mg/kg. Network pharmacological research analysis identified 91 putative rutin targets associated with IS, including key inflammatory mediators such as interleukin - 6 (IL-6), tumor necrosis factor-α (TNF-α) and IL-1β. After rutin intervention, the levels of nuclear factor kappa-B (NF-κB) p65, NOD-like Receptor thermal protein domain associated protein 3 (NLRP3), Caspase-1, IL-1β, IL-18, IL-6, and TNF-α were significantly reduced, while Gastermin D (GMDSD) was increased in tMCAO rats. In the OGD/R model, rutin inhibited the activation of NLRP3, and further enhanced the inhibition of NF-κB in combination with JSH-23.

Conclusions: This study delineates a multimodal neuroprotective efficacy of rutin in IS, mechanistically characterized by its coordinated suppression of the neuroinflammation-pyroptosis axis via NF-κB/NLRP3 pathway modulation. The dose-dependent attenuation of astrocytic homeostasis disruption further substantiates its therapeutic precision, consistent with the multitarget engagement patterns predicted by network pharmacology. Notably, functional validation with an NLRP3 Agonist highlights pathway specificity, positioning rutin as a promising phytochemical scaffold for the development of neurovascular unit-stabilizing agents. These findings not only broaden the pharmacodynamic paradigm of flavonoid-based stroke therapeutics but also advocate for systematic exploration of rutin's translational potential in modulating neuroinflammatory cascades across neurological disorders.

Ischemic stroke; NF-κB; NLRP3; Neuroinflammation; Rutin.