Low-cytotoxic synthetic bromorutaecarpine exhibits anti-inflammation and activation of transient receptor potential vanilloid type 1 activities

Abstract

Rutaecarpine (RUT), the major bioactive ingredient isolated from the Chinese herb Evodia rutaecarpa, possesses a wide spectrum of biological activities, including anti-inflammation and preventing cardiovascular diseases. However, its high cytotoxicity hampers pharmaceutical development. We designed and synthesized a derivative of RUT, bromo-dimethoxyrutaecarpine (Br-RUT), which showed no cytotoxicity at 20 μM. Br-RUT suppressed nitric oxide (NO) production and tumor necrosis factor-α release in concentration-dependent (0~20 μM) manners in lipopolysaccharide (LPS)-treated RAW 264.7 macrophages; protein levels of inducible NO synthase (iNOS) and cyclooxygenase-2 induced by LPS were downregulated. Br-RUT inhibited cell migration and invasion of ovarian carcinoma A2780 cells with 0~48 h of treatment. Furthermore, Br-RUT enhanced the expression of transient receptor potential vanilloid type 1 and activated endothelial NOS in human aortic endothelial cells. These results suggest that the synthetic Br-RUT possesses very low cytotoxicity but retains its activities against inflammation and vasodilation that could be beneficial for cardiovascular disease therapeutics.

Scheme 1

Synthesis of bromo-dimethoxyrutaecarpine (Br-RUT).

Figure 1

Effects of bromo-dimethoxyrutaecarpine (Br-RUT) on nitric oxide (NO) and tumor necrosis factor (TNF)-α releases by lipopolysaccharide (LPS)-treated (100 ng/mL) RAW 264.7 macrophages. (a) NO levels were detected in culture medium using the Griess reaction. The percentage of untreated cells was set as the control to 100%. (b) TNF-α release in cell supernatants was detected using a mouse TNF-α Quantikine kit. (c) Cell viability upon Br-BUT treatment for 24 h in an MTT assay. Statistical significance is indicated compared to LPS treatment. (*P < 0.05, **P < 0.001, ***P < 0.001).

Figure 2

Figure 3

Effects of bromo-dimethoxyrutaecarpine (Br-RUT) on cell migration and invasion. Cell migration (a) and invasion (b) were detected following Br-RUT treatment for 0~48 h and photographed with a microscope (upper panel). Stained cells were counted and calculated in three random regions for each sample, results are presented as the mean ± SD from triplicate experiments, and the statistical analysis is shown in the lower panel.

Figure 4

Effects of bromo-dimethoxyrutaecarpine (Br-RUT) on transient receptor potential vanilloid type 1 (TRPV1) expression and endothelial nitric oxide synthase (eNOS) phosphorylation in human aortic endothelial cells (HAECs). The densitometric ratio is indicated.