METHODS: Time-kill analysis of one MRSA reference strain (ATCC 43300) and three clinical isolates (WM3, BM1 and KJ7) for both compounds was first performed to provide the bacteriostatic/bactericidal profile. Then, MRSA ATCC 43300 strain treated with both compounds was interrogated by NGS.
RESULTS: Both stigmasterol and lupeol possessed bacteriostatic properties against all MRSA tested; however, lupeol exhibited both bacteriostatic and bactericidal properties within the same minimum inhibitory concentration and minimum bactericidal concentration values against BM1 (12.5mg/mL). Transcriptome profiling of MRSA ATCC 43300 revealed significant modulation of gene expression with multiple desirable targets by both compounds, which caused a reduction in the translation processes leading to inhibition of protein synthesis and prevention of bacterial growth.
CONCLUSIONS: This study highlights the potential of both stigmasterol and lupeol as new promising anti-MRSA agents.
METHODOLOGY/PRINCIPAL FINDINGS: PA exerted selective cytotoxicity on human umbilical vein endothelial cells (HUVECs) with IC(50) value of 6.91 ± 0.85 µM when compared to human normal fibroblast and normal liver epithelial cells. Assessment of the growth kinetics by cell impedance-based Real-Time Cell Analyzer showed that PA induced both cytotoxic and cytostatic effects on HUVECs, depending on the concentration used. Results also showed that PA suppressed VEGF-induced survival and proliferation of HUVECs. Furthermore, endothelial cell migration, invasion, and morphogenesis or tube formation demonstrated significant time- and dose-dependent inhibition by PA. PA also suppressed matrix metalloproteinase-2 (MMP-2) secretion and attenuated its activation to intermediate and active MMP-2. In addition, PA suppressed F-actin stress fiber formation to prevent migration of the endothelial cells. More importantly, anti-angiogenic potential of PA was also evidenced in two in vivo models. PA inhibited neo-vessels formation in murine Matrigel plugs, and angiogenesis in zebrafish embryos.
CONCLUSIONS/SIGNIFICANCE: Taken together, our study demonstrated the distinctive anti-angiogenic properties of PA, both in vitro and in vivo. This report thus reveals another biological activity of PA in addition to its reported anti-inflammatory and anti-cancer activities, suggestive of PA's potential for development as an anti-angiogenic agent for cancer therapy.