背景:天然来源的化合物是具有独特作用机制的有效药物来源。在植物化学物质中,反式肉桂醛(t-CA)具有广泛的生物活性,因此,几个世纪以来一直被用来对抗细菌和真菌感染。然而,这些性质的分子基础还没有被完全覆盖。考虑到难以控制的感染正在成为一个日益严重的全球问题,有必要阐明t-CA的分子潜力。
目的:通过抑制毒力因子的表达,评价t-CA对志贺产毒大肠杆菌的抗菌活性,阐明其作用机制。
方法:t-CA的抗微生物潜力用两倍微量稀释和时间-杀灭试验进行评估。进一步的评估包括生物发光抑制试验,活性氧(ROS)的定量和NAD+/NADH比率的评估。使用透射电子显微镜检查t-CA暴露后的形态变化。RNA测序和核苷酸的放射性标记阐明了由t-CA诱导的代谢改变。通过应用融合蛋白监测毒素表达水平,监测噬菌体的发展,和荧光显微镜研究。最后,使用Galleriamellonella感染模型评估体内治疗效果。
结果:对t-CA生物活性的全面研究显示了影响细菌代谢和形态的独特特性,导致显著的细菌细胞变形和有效的毒力抑制。对潜在机制的阐明表明,t-CA激活了全球监管体系,严厉的回应,表现为它的警报,(p)ppGpp,RelA酶介导的过度生产,从而抑制细菌增殖。有趣的是,t-CA通过Alarmone分子有效下调志贺毒素基因表达,表明其潜在的治疗效果。体内验证表明,在t-CA治疗后,使用50mg/kg(p<0.05),幼虫存活率显着提高,类似于阿奇霉素观察到的疗效,因此表明其对EHEC感染的有效性(p<0.05)。
结论:总的来说,这些结果揭示了t-CA强大的抗菌能力,保证其作为一种可行的抗感染剂的进一步探索。
BACKGROUND: Compounds of natural origin are potent source of drugs with unique mechanisms of action. Among phytochemicals, trans-cinnamaldehyde (t-CA) exhibits a wide range of biological activity, thus has been used for centuries to fight bacterial and fungal infections. However, the molecular basis of these properties has not been fully covered. Considering that difficult-to-control infections are becoming a rising global problem, there is a need to elucidate the molecular potential of t-CA.
OBJECTIVE: To evaluate the antibacterial activity of t-CA against Shiga-toxigenic E. coli strains and elucidate its mechanism of action based on the inhibition of the virulence factor expression.
METHODS: The antimicrobial potential of t-CA was assessed with two-fold microdilution and time-kill assays. Further evaluation included bioluminescence suppression assays, quantification of reactive oxygen species (ROS) and assessment of NAD+/NADH ratios. Morphological changes post t-CA exposure were examined using transmission electron microscopy. RNA sequencing and radiolabeling of nucleotides elucidated the metabolic alterations induced by t-CA. Toxin expression level was monitored through the application of fusion proteins, monitoring of bacteriophage development, and fluorescence microscopy studies. Lastly, the therapeutic efficacy in vivo was assessed using Galleria mellonella infection model.
RESULTS: A comprehensive study of t-CA\'s bioactivity showed unique properties affecting bacterial metabolism and morphology, resulting in significant bacterial cell deformation and effective virulence inhibition. Elucidation of the underlying mechanisms indicated that t-CA activates the global regulatory system, the stringent response, manifested by its alarmone, (p)ppGpp, overproduction mediated by the RelA enzyme, thereby inhibiting bacterial proliferation. Intriguingly, t-CA effectively downregulates Shiga toxin gene expression via alarmone molecules, indicating its potential for therapeutic effect. In vivo validation demonstrated a significant improvement in larval survival rates post- t-CA treatment with 50 mg/kg (p < 0.05), akin to the efficacy observed with azithromycin, thus indicating its effectiveness against EHEC infections (p < 0.05).
CONCLUSIONS: Collectively, these results reveal the robust antibacterial capabilities of t-CA, warranting its further exploration as a viable anti-infective agent.