Flavonoid-rich Phyla nodiflora fraction promotes Keap1 degradation and Nrf2/HO-1 activation to attenuate particulate matter-induced oxidative stress in human keratinocytes

J Ethnopharmacol. 2026 Apr 6:360:121168. doi: 10.1016/j.jep.2026.121168. Epub 2026 Jan 4.

Abstract

Ethnopharmacological relevance: Phyla nodiflora (L.) Greene (PN) has been traditionally used in Asia for the treatment of inflammatory and dermatological conditions. Its pharmacological activities are largely attributed to flavonoids with antioxidant and anti-melanogenic effects. Given these traditional and pharmacological properties, it is plausible that PN may protect the skin cells against airborne stressors, such as particulate matter (PM).

Aim of the study: To evaluate a flavonoid-rich fraction of PN derived from the solvent-free microwave hydrodiffusion and gravity extraction (PN Fr. 5) for its protective effects against PM-induced oxidative damage.

Materials and methods: Human keratinocytes (HaCaT cells) were exposed to urban dust PM (NIST® SRM® 1649b). PN antioxidant activity was evaluated by measuring PM-induced intracellular reactive oxygen species (ROS), and mechanisms were examined via the Keap1-Nrf2/HO-1 pathway using Western blotting, RT-qPCR, and cycloheximide chase assays. Feature-based molecular networking and docking were used to prioritize putative active compounds.

Results: PN Fr. 5 showed the strongest activity among preparations, reducing PM-induced ROS and upregulating AQP3 without cytotoxicity. Mechanistically, PN Fr. 5 accelerated Keap1 degradation, stabilizing Nrf2 and upregulating downstream antioxidant proteins, including HO-1. Molecular networking identified jaceosidin, isokaempferide, eupafolin, and chrysosplenol D as the major flavonoids, with docking studies suggesting that these compounds, particularly jaceosidin, may collectively contribute as non-electrophilic modulators of the Keap1-Nrf2 protein-protein interaction, thereby enhancing Nrf2/HO-1 signaling.

Conclusions: The flavonoid-rich fraction of P. nodiflora (PN Fr. 5) exhibited potent antioxidant activity through Keap1-Nrf2/HO-1 activation, supporting its traditional dermatological use and potential as a plant-based protective agent against PM-induced oxidative damage in keratinocytes.

Keywords: Flavonoids; Keap1–Nrf2/HO-1 pathway; Keratinocyte; Particulate matter; Phyla nodiflora.

MeSH terms

  • Antioxidants* / isolation & purification
  • Antioxidants* / pharmacology
  • Cell Line
  • Flavonoids* / isolation & purification
  • Flavonoids* / pharmacology
  • HaCaT Cells
  • Heme Oxygenase-1 / metabolism
  • Humans
  • Kelch-Like ECH-Associated Protein 1* / metabolism
  • Keratinocytes* / drug effects
  • Keratinocytes* / metabolism
  • Molecular Docking Simulation
  • NF-E2-Related Factor 2* / metabolism
  • Oxidative Stress* / drug effects
  • Particulate Matter* / toxicity
  • Plant Extracts* / pharmacology
  • Reactive Oxygen Species / metabolism
  • Signal Transduction / drug effects

Substances

  • NF-E2-Related Factor 2
  • Kelch-Like ECH-Associated Protein 1
  • Flavonoids
  • Particulate Matter
  • NFE2L2 protein, human
  • KEAP1 protein, human
  • Plant Extracts
  • Antioxidants
  • Heme Oxygenase-1
  • HMOX1 protein, human
  • Reactive Oxygen Species