CHR抑制白假丝酵母菌生物被膜形成作用及机制探讨

Inhibiting effect of CHR on biofilm formation of Candida albicans and its mechanism

目的：CGA47-66（chromofungin,CHR）是嗜铬粒蛋白A（chromograinin A,CGA）水解形成的生物活性多肽,已有研究证实其具有免疫调节和抗菌作用,本研究旨在进一步证实CHR抗白假丝酵母菌生物被膜形成的作用,探讨其作用机制。方法：根据CLSI-M27-A3参考方法检测CHR对浮游状态下白假丝酵母菌的最低抑菌浓度,菌落计数法及XTT比色法验证CHR是否具有抑制生物被膜形成的作用,并确定CHR对白假丝酵母菌的BIC50,通过菌落计数法和倒置显微镜证实CHR抗真菌粘附作用,并使用q RT-PCR检测粘附基因Hwp1的表达情况从而进一步验证其作用的分子机制。结果：CHR对浮游状态白假丝酵母菌的MIC值为10～20μmol/L,并证实CHR具有抗白假丝酵母菌生物被膜形成作用,BIC50值为80～160μmol/L,且具有浓度依赖性;q RT-PCR检测CHR下调C.albicans Hwp1基因的表达,160μmol/L CHR处理组相对Hwp1基因表达量通过2-△△Ct值计算为（0.089 5±0.036 0）,无处理组Hwp1基因的表达是CHR组的11.17倍,差异具有统计学意义（t=44.008,P=0.001）。结论：CHR具有抑制白假丝酵母菌生物被膜形成的作用,其机制可能与CHR减少白假丝酵母菌细胞的粘附有关。本研究为CHR在抗真菌材料方面的应用提供了理论基础。

Objective：To further confirm if CGA47-66（chromofungin,CHR）plays an inhibitory role on Candida albicans biofilm formation,and to explore its possible mechanism. Methods：MIC50of CHR on floating state Candida albicans was detected by M27-A3 reference method. BIC50 of CHR on biofilm formation was observed by XTT colorimetric assay;colony counting method and XTT colorimetric assay were employed to validate whether CHR had an inhibitory function on Candida albicans biofilm formation,then the antifungal adhesion function of CHR was tested by colony counting method combined with inverted microscope. The further molecular mechanism,and the expressions of adhesion gene Hwp1 were measured by q RT-PCR. Results：Our study firstly demonstrated that CHR could inhibit biofilm formation in a dose dependent manner;the BIC50 was 80 to 160 μmol/L;the MIC50 value of CHR on floating state Candida albicans was 10 to 20μmol/L. CHR could down regulate the expression of adhesion gene Hwp1. The relative expression fold of CHR group was（0.089 5±0.036 0）,the expression of gene Hwp1 in control group was11.17 times than that in the CHR group,and the difference had statistical significance（t=44.008,P=0.001）. Conclusion：CHR has the inhibitory effect on Candida albicans biofilm formation,the mechanism may related to reducing adherent ability of Candida albicans.This study provides theoretical basis for the application of CHR in the field of antifungal materials.