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目的:探究氟康唑(FLC)和他克莫司(FK506)联用对光滑念珠菌耐药性及CDR1、ABC转运蛋白表达的影响。方法:选择临床分离的耐药性光滑念珠菌,采用体外微量稀释法测量FLC和FK506联用对光滑念珠菌的体外静态抗真菌作用,选择部分抑菌浓度指数和FICI法对结果进行评价分析。采用RT-PCR法检测药物联合干预后光滑念珠菌耐药基因CDR1的相对表达量,通过测定菌株细胞外液FDA浓度来对ABC转运蛋白的外排情况进行间接观察。结果:FK506的加入使得对FLC耐药光滑念珠菌的MIC值在24 h作用时降低至原来的1/8,在48 h作用时降低至原来的1/16,FLC和FK506联合用药抗光滑念珠菌在24 h和48 h作用的FICI值分别为0.089、0.124,均<0.5,表明FLC与FK506联用对光滑念珠菌具有较强的协同作用。与未加药组相比,FLC与FK506单独用药后CDR1基因的表达水平无明显增加(P>0.05),FDA的细胞外液浓度升高水平降低(P<0.05),且FLC与FK506联合用药后FDA细胞外液浓度的升高水平低于各自单独用药后,差异具有统计学意义(P<0.05)。结论:FLC与FK506联用能协同抗光滑念珠菌,有效克服其对FLC的耐药性,其机制可能与降低CDR1基因及ABC转运蛋白的表达量有关。
Objective: To investigate the effects of fluconazole (FLC) and tacrolimus (FK506) on the drug resistance of Candida glabrata and the expression of CDR1 and ABC transporter. Methods: The clinical isolates of Candida glabrata were selected. The in vitro antifungal activity of FLC and FK506 against Candida glabrata was measured by in vitro microdilution method. The results of partial antibacterial concentration index and FICI method were evaluated. The relative expression level of Candida glabrata-resistant gene CDR1 was detected by RT-PCR. The efflux of ABC transporter was observed indirectly by measuring the concentration of extracellular fluid FDA. RESULTS: The addition of FK506 reduced the MIC of FLC-resistant Candida glabrata to 1/8 at 24 h and to 1/16 at 48 h. FLC and FK506 combination anti-smooth rosary beads The FICI values of bacteria at 24 h and 48 h were 0.089 and 0.124, respectively, which were all <0.5, indicating that FLC combined with FK506 had a strong synergistic effect on Candida glabrata. Compared with the untreated group, the expression of CDR1 gene in FLC group and FK506 alone group had no significant increase (P> 0.05), and the level of FDA extracellular fluid concentration decreased (P <0.05), and the combined use of FLC and FK506 Post-FDA levels of extracellular fluid concentrations were lower than the respective drug alone, the difference was statistically significant (P <0.05). Conclusion: FLC combined with FK506 can synergize with Candida glabrata to effectively overcome its resistance to FLC, and its mechanism may be related to the decrease of CDR1 gene and ABC transporter expression.