Cardiospermum halicacabum Extract Inhibits the Production of Nitric Oxide and Reactive Oxygen Species in LPS-Stimulated RAW 264.7 Cells through Erk/p38 Signalling Pathways

The medicinal plant Cardiospermum halicacabum has been extensively studied for its potential in various therapeutic activities, including anti-inflammation, antioxidant, antibacterial, antifungal and antiparasitic effects. However, the specific molecular mechanisms responsible for its anti-inflammatory properties remain largely unknown. In order to shed light on this aspect, the current investigation was aimed at elucidating the underlying mechanisms through which C. halicacabum exerts its anti-inflammatory response. The different concentrations of C. halicacabum extract were tested for cytotoxicity on RAW264.6 murine macrophage cells by MTT assay. The inhibition of nitric oxide (NO) in LPS-induced cell pre-treated with the extract was evaluated by Griess assay using Griess reagent. Flow cytometry and fluorescent microscopy were used to examine the reactive oxygen species (ROS) in the treated cells. The expression of iNOS and the phosphorylation status of Erk1/2, p38 and NF-kB were determined by western blotting. The C. halicacabum extract at concentrations up to 125 µg/mL did not cause the cytotoxicity to the RAW264.6 murine macrophage cells. Incubation of LPS-induced cells with the extract significantly minimized the production of NO as compared with the LPS-stimulated cells. Further, the extract significantly reduced ROS generation in LPS-induced cells. In the present of C. halicacabum, LPS-treated cells had the reduced NOS expression as well as had low phosphorylation on Erk1/2, p38 and NF-kB. C. halicacabum attenuates NO and ROS production under the regulation of Erk1/2 and p38 pathway as well as NF-kB. HIGHLIGHTS The underlying mechanisms of halicacabum on its anti-inflammatory response were investigated. halicacabum extract significantly reduced ROS and NO generation in LPS-induced cells. halicacabum attenuates NO and ROS production under the regulation of Erk1/2 and p38 pathway as well as NF-kB. GRAPHICAL ABSTRACT