应用角质层真菌计量生物学分析法建立药敏试验新模型

A new bioassay model of antibiotic susceptibility by corneofungimetry

目的应用角质层真菌计量生物学分析法建立药敏试验新模型,以更好地评价抗真菌药物的体内外一致性。方法21名健康志愿者被随机分为两组,分别口服不同剂量(200mg,2次/d;200mg,1次/d)的伊曲康唑1周;取志愿者服药前、服药后不同时间(多次)皮肤角质层作为培养基(皮肤角质层剥离条),将红色毛癣菌、须毛癣菌、犬小孢子菌、白念珠菌、光滑念珠菌和热带念珠菌的菌悬液分别接种其上。培养后,行碘酸-雪夫(PAS)染色;采用角质层真菌计量生物学分析法并辅以计算机辅助图像分析,测定被孢子和菌丝覆盖的面积大小。结果在两组受试者的角质层培养基上6种真菌均可生长,但随着药物在角质层中的含量(浓度)不同,菌落的大小、菌丝稠密度、孢子数量也不同。真菌的生长受到药物的抑制,尤其是服药7d后真菌覆盖的面积明显低于服药前(P<0·01)。两种不同服药方法对3种皮肤癣菌及白念珠菌和热带念珠菌的抑制作用差异无统计学意义,均P>0.05。而对于光滑念珠菌差异则有统计学意义,P<0.01。伊曲康唑在人体角质层的生物利用度较高,对6株菌表现了很高的抗真菌活性。伊曲康唑在人体角质层的生物利用度较高,对6株菌表现了很高的抗真菌活性。结论应用改良的角质层真菌计量生物学分析辅以计算机图形分析法进行药物敏感性分析在概念上有别于传统的体外药敏试验,建立了一种与体内环境相似的新模型,对评价抗皮肤真菌药物的体内外一致性有一定的参考价值。

Objective To study the value of a new bioassay model of antibiotic susceptibility by corneofungimetry which is more similar to human internal environment. Methods Twenty-one healthy adult volunteers, 12 males and 9 females, aged 32 （22-41 ）, were randomly divided into two groups to receive itraconazole 200 mg Bid or 200 mg qd or bid for one week. Stratum corneum strippings were taken from the forearm skin once before administration of itraconazole and 1, 4, 7, 10, 14, 21, 28, and 35 days after administration of itraconazole. Spores of selected fungi （Trichophyton rubrum, T. mentagrophytes, Microsporum canis, Candida albicans, C. glabrata and C. tropicalis ） were deposited and cultured on the strippings. PAS stain was used after one-week culture to observe the growth of the fungi by computerizedassisted image analysis to measure the area covered by the yeasts and the dermatophyte. Results All fungi grew on all the stratum corneum strippings with different numbers of hyphae. The areas covered by hyphae and spores decreased gradually after administration of itraconazole, became the smaller on the seventh day （P = 0.01 or P 〈 0.01 ）, and began to increased since the 10 th day, and became almost the same sizes as those before the administration of itraconazole on the 35 th day. Conclusion Similar to the human internal environment, corneofungimetry is a useful new model of bioassay for antibiotic susceptibility.