真菌性角膜炎病原学及与TLR2/MyD88/NF-κB p65信号通路的关联

Etiological characteristics of fungal keratitis and its association with TLR2/MyD88/NF-κB p65 signaling pathway

目的调查真菌性角膜炎(FK)病原学分布特征和耐药性,同时分析角膜上皮细胞Toll样受体2(TLR2)基因多态性及表达水平。方法选取2018年1月-2021年12月于河南省人民医院接受诊治的85例FK患者为样本进行横断面研究,刮取病灶边缘角膜组织,鉴定病原菌分布和耐药性,检测TLR2基因rs380409位点单核苷酸多态性(SNP)以及TLR2、髓样分化因子88(MyD88)、核因子κB(NF-κB)p65表达水平,分析FK患者TLR2/MyD88/NF-κB p65信号通路分子表达水平与真菌类型、病情严重程度和TLR2基因rs380409位点SNP的关系。结果85例FK患者共检出真菌97株,其中镰刀菌属、曲霉菌属和假丝酵母属占比分别为31.96%、26.80%和19.59%;镰刀菌属耐药率较低,曲霉菌属对两性霉素B和伏立康唑耐药率较低,对伊曲康唑和氟康唑耐药性较高,假丝酵母属对两性霉素B耐药率较低,对伏立康唑、伊曲康唑和氟康唑耐药性较高;轻度、中度和重度FK患者角膜上皮TLR2、MyD88和NF-κB p65表达水平上升(P<0.05),各组比较差异有统计学意义(P<0.05);85例FK患者TLR2 rs3804099基因型分别为TT型31例(36.47%)、TC型35例(41.18%)和CC型19例(22.35%),Hardy-Weinberg法检验显示基因型分布满足平衡规律;TLR2基因rs3804099位点TT和TC基因型患者角膜组织TLR2、MyD88和NF-κB p65表达水平均低于CC基因型(P<0.05)。结论镰刀菌属、曲霉菌属和假丝酵母属为FK常见致病菌,对两性霉素B耐药性较低,对伏立康唑、伊曲康唑和氟康唑耐药性较高,此外FK发病和进展与TLR2/MyD88/NF-κB信号通路激活密切相关。

OBJECTIVE To investigate the etiological distribution characteristics and drug resistance in fungal keratitis(FK),and to analyze the gene polymorphism and expression level of corneal epithelial cell Toll-like receptor 2(TLR2).METHODS A cross-sectional study was conducted on 85 patients with FK who were diagnosed and treated in Henan Provincial People′s Hospital between Jan 2018 and Dec 2021.The marginal corneal tissues of the lesions were scraped to identify the distribution and drug resistance of pathogenic bacteria,and the single nucleotide polymorphism(SNP)at rs3804099 locus of TLR2 gene and expression levels of TLR2,myeloid differentiation factor 88(MyD88)and nuclear factorκB(NF-κB)p65 were detected.The relationship between expression levels of TLR2/MyD88/NF-κB p65 signaling pathway molecules and fungal type,disease severity and SNP at rs3804099 locus of TLR2 gene were analyzed among patients with FK.RESULTS A total of 97 strains of fungi were detected among 85 patients with FK,of which Fusarium,Aspergillus and Candida accounted for 31.96%,26.80%and 19.59%,respectively.Fusarium had low drug-resistant rate;Aspergillus was lowly drug-resistant to amphotericin B and voriconazole but highly resistant to itraconazole and fluconazole;Candida was lowly drug-resistant to amphotericin B but highly drug-resistant to voriconazole,itraconazole and fluconazole.The expression levels of TLR2,MyD88 and NF-κB p65 in corneal epithelium of patients with mild,moderate and severe FK showed upward trends(P<0.05),and the differences between the groups were statistically significant(P<0.05).The genotypes at rs3804099 locus of TLR2 in 85 patients with FK were TT genotype in 31 cases(36.47%),TC genotype in 35 cases(41.18%)and CC genotype in 19 cases(22.35%).Hardy-Weinberg test showed that the genotype distribution satisfied the law of equilibrium.The expression levels of TLR2,MyD88 and NF-κB p65 in corneal tissue of patients carrying TT genotype or TC genotype at rs3804099 locus of TLR2 gene were lower than those carrying CC genotype(P<0.05).CONCLUSION Fusarium,Aspergillus and Candida sorokinanum are common pathogenic bacteria of FK,with low drug resistance to amphotericin B but high drug resistance to voriconazole,itraconazole and fluconazole.The onset and progression of FK are closely related to the activation of TLR2/MyD88/NF-κB signaling pathway.