法尼醇对白念珠菌生物膜死亡作用的研究初探

A preliminary study of the effect of farnesol on cell death in C.albicans biofilms

目的探讨法尼醇对白念珠菌生物膜死亡的作用是坏死作用还是凋亡作用,并初步探讨其与CDR1基因及MCA1基因表达的相关性。方法研究菌株为白念珠菌标准株和耐药株,研究分为白念珠菌生物膜法尼醇处理组及未处理对照组。采用激光共聚焦显微镜观察PI荧光染色后法尼醇对生物膜细胞坏死的形态学变化。采用Annexin V-FITC/PI流式分析法尼醇对白念珠菌生物膜凋亡和坏死的作用,q-PCR分析法尼醇对白念珠菌坏死和凋亡相关基因CDR1及MCA1表达的改变。结果法尼醇处理组白念珠菌细胞坏死率高于未处理对照组(P<0.05)。在不同生物膜时相,法尼醇对耐药株和标准株的坏死作用具有一致性。在400~800μmol/L法尼醇处理下,6 h生物膜坏死率低于12 h生物膜,高于24 h生物膜(P<0.05)。此外,法尼醇对耐药株生物膜作用后的坏死率高于标准株(P<0.05)。法尼醇对白念珠菌耐药株和标准株凋亡作用不明显(P>0.05)。法尼醇增高耐药株各时相及标准株早期生物膜CDR1基因表达,且增高耐药株早期和标准株早期生物膜的MCA1基因表达(P<0.05)。结论法尼醇对白念珠菌耐药株和标准株生物膜的死亡作用以促进坏死为主,且对耐药株的坏死作用较对标准株显著。CDR1基因和MCA1基因与法尼醇对耐药株生物膜的坏死作用相关。法尼醇对白念珠菌生物膜的坏死作用与其浓度和生物膜时相相关。

Objective To investigate the effect of farnesol on cell death in C.albicans biofilms is necrosis or apoptosis, and primarily discuss its correlation with the expression of CDR1 gene and MCA1 gene. Methods The research strains were C.albicans standard strain and resistant strain. The study was divided into C.albicans biofilm farnesol treated group and its untreated control group. Morphological changes of C.albicans biofilm treated with farnesol were observed by confocal laser scanning microscope with PI staining. Annexin V-FITC/PI flow cytometry was employed to analyze the effect of farnesol on apoptosis and necrosis of C.albicans. The expression changes of CDR1 and MCA1 gene related to apoptosis and necrosis of C.albicans were detected through q-RTPCR. Results The study found that the necrotic rate of C.albicans biofilms was higher in farnesol treated groups than that of the non-treated groups(P<0.029). In different biofilm phases, farnesol had a consistent necrosis effect on the biofilms of the resistant and standard strains. Under the treatment of 400-800 μmol/L farnesol, the rate of necrosis cells of 6 h phase was lower than 12 h phase, but higher than 24 h phase(P<0.021). In addition, the rate of necrosis cells was higher in biofilms of the resistant strains than that of the biofilms of the standard strains(P<0.002). The apoptosis effect of farnesol on C.albicans biofilms of the resistant and standard strains was not obvious(P>0.05). Farnesol increased the expression of CDR1 genes of the biofilms of the resistant strains at three studied phases and that of the biofilms of the standard strains at the early phase, respectively. Farnesol also increased the expression of MCA1 gene of the biofilms of the resistant strains and standard strains at the early phase, respectively(P<0.023). Conclusion Farnesol promotes the necrosis of C.albicans biofilms of the resistant and standard strains, which is more significant on the biofilms of the resistant strains. CDR1 gene and MCA1 gene may be associated with necrosis of biofilms formed from the resistant strain. The necrosis effect of farnesol on the biofilm of C.albicans is related to its concentration and biofilm phase.